[Title 32 CFR 627]
[Code of Federal Regulations (annual edition) - July 1, 2002 Edition]
[Title 32 - NATIONAL DEFENSE]
[Chapter V - DEPARTMENT OF THE ARMY]
[Subchapter H - SUPPLIES AND EQUIPMENT]
[Part 627 - THE BIOLOGICAL DEFENSE SAFETY PROGRAM, TECHNICAL SAFETY REQUIREMENTS (DA PAMPHLET 385-69)]
[From the U.S. Government Printing Office]
32NATIONAL DEFENSE32002-07-012002-07-01falseTHE BIOLOGICAL DEFENSE SAFETY PROGRAM, TECHNICAL SAFETY REQUIREMENTS (DA PAMPHLET 385-69)627PART 627NATIONAL DEFENSEDEPARTMENT OF THE ARMYSUPPLIES AND EQUIPMENT
PART 627--THE BIOLOGICAL DEFENSE SAFETY PROGRAM, TECHNICAL SAFETY REQUIREMENTS (DA PAMPHLET 385-69)--Table of Contents
Subpart A--Introduction
Sec.
627.1 Purpose.
627.2 Background.
627.3 Scope.
627.4 References.
627.5 Abbreviations and terms.
Subpart B--Administration
627.6 Safety administration.
627.7 Goal of a laboratory safety program.
627.8 Occupational health.
627.9 Medical records.
Subpart C--Operational Requirements
627.10 Personnel prerequisites.
627.11 Operational prerequisites.
627.12 General laboratory techniques.
627.13 Biosafety level 1.
627.14 Biosafety level 2.
627.15 Biosafety level 3.
627.16 Biosafety level 4.
627.17 Toxins.
627.18 Emergencies.
627.19 Large-scale operations.
627.20 Operations with radioactive material.
Subpart D--Personal Protective Equipment
627.21 Introduction.
627.22 Minimum laboratory attire for use of etiologic agents.
627.23 Biosafety level 1.
627.24 Biosafety level 2.
627.25 Biosafety level 3.
627.26 Biosafety level 4.
627.27 Large-scale (LS) operations.
627.28 Solutions of toxins and dry forms of toxins in closed
containers.
627.29 Dry forms of toxins handled in open containers.
627.30 Situations specified in Sec. 627.18(e).
627.31 Specific requirements for individual PPE items.
Subpart E--Decontamination and Disposal
627.32 Introduction.
627.33 Methods of decontamination.
627.34 Disposal.
Subpart F--Importation, Shipment, and Transport of Etiologic Agents
627.35 Introduction.
627.36 Administration.
627.37 Importation directives.
627.38 Shipment directives.
627.39 Transportation directives.
627.40 Additional requirements.
627.41 Sources for further information on shipment of etiologic agents.
Subpart G--Facilities
627.42 Introduction.
627.43 Biosafety level 1.
627.44 Biosafety level 2.
627.45 Biosafety level 3.
627.46 Biosafety level 4.
627.47 Large-scale facilities.
627.48 Toxins.
Subpart H--Engineering Controls
627.49 Introduction.
627.50 Class I biological safety cabinet.
627.51 Class II biological safety cabinet.
627.52 Class III biological safety cabinet.
627.53 Fume hood.
627.54 Glove box.
627.55 Ventilated balance enclosures.
627.56 Ventilated cage enclosures.
627.57 Ventilated cage areas.
Appendix A to Part 627--References
Appendix B to Part 627--Resource List for Immunoprophylaxis of Personnel
at Risk
Appendix C to Part 627--Laboratory Safety Inspection Checklist
Appendix D to Part 627--Packaging and Labeling Requirements for Shipment
of Etiologic Agents
Appendix E to Part 627--Permits for Importation and Shipment of
Etiologic Agents
Appendix F to Part 627--Drawings, Biological Safety Cabinets
Appendix G to Part 627--Glossary
[[Page 534]]
Authority: 5 U.S.C. 102, 21 U.S.C. 111, 151-158; 42 U.S.C. 216; sec.
361, 58 Stat. 703 and 264; 49 U.S.C. App. 1803, 1804, 1807, and 1808; 50
U.S.C. 1431, 29 CFR 1910. 1450(e) and Public Law 101-510, 104 Stat.
1516.
Source: 57 FR 12604, Apr. 10, 1992, unless otherwise noted.
Subpart A--Introduction
Sec. 627.1 Purpose.
This pamphlet prescribes the technical safety requirements for the
use, handling, shipment, storage, and disposal of etiologic agents used
in research, development, test, and evaluation (RDTE) for the Biological
Defense Program (BDP)
Sec. 627.2 Background.
The United States Army BDP, on behalf of the Department of Defense,
supports RDTE efforts to maintain and develop defensive measures and
materiel to meet potential biological warfare threats. The program's
objectives are to develop measures for identification, detection,
treatment, protection against, and decontamination of these threats. To
meet the program objectives, etiologic agents are used to conduct the
necessary handling, storage, shipment, and disposal of etiologic agents.
This pamphlet describes requirements based on Centers for Disease
Control-National Institute of Health (CDC) (NIH) guidelines, Biosafety
in Microbiological and Biomedical Laboratories, and establishes
guidelines for toxins.
Sec. 627.3 Scope.
The requirements stated in this pamphlet apply to all elements of
the Army to include the ARNG and the USAR and its contractors and
subcontractors who use, produce, store, handle, or ship etiologic agents
in support of the BDP, regardless of the source of the agent(s).
Sec. 627.4 References.
Required and related publications are listed in appendix A of this
part.
Sec. 627.5 Abbreviations and terms.
Abbreviations and special terms used in this part are explained in
appendix F of this part.
Subpart B--Administration
Sec. 627.6 Safety administration.
Each BDP institution must have a safety program that complies with
AR 385-10, AR 385-69, and this pamphlet. In addition, the safety program
must be designed to ensure compliance with--
(a) Occupational Safety and Health Administration (OSHA)
requirements for health and safety.
(b) Environmental Protection Agency (EPA) regulations designed to
implement the Resource Conservation and Recovery Act (RCRA) and the
National Environmental Policy Act (NEPA).
(c) Nuclear Regulatory Commission (NRC) requirements for safe
handling of radioactive isotopes (when applicable).
(d) NIH Guidelines for Research Involving Recombinant
Deoxyribonucleic Acid (DNA) Molecules.
(e) Relevant national, State, and local regulations.
(f) Any requirements of applicable accrediting bodies.
Sec. 627.7 Goal of a laboratory safety program.
The goals of the laboratory safety program are to protect those
working in the laboratory, others who may potentially be exposed to
hazards in the laboratory, and the environment. In addition, a
laboratory safety program should ensure that hazardous materials will be
handled and disposed of in such a way that people, other living
organisms, and the environment are protected from harm. Safety awareness
must be a part of everyone's habits, and can only be achieved if all
senior and responsible staff have a sincere, visible, and continuing
interest in preventing injuries and occupational illnesses. Laboratory
personnel, for their part, must carry out their work in a way that
protects themselves and their fellow workers.
(a) Laboratory safety. The safety program will be carried out as
stated in AR 385-69. Additionally, the program will contain the
following elements--
(1) The commander or institute director, along with all personnel,
must have a continuing, observable, and known commitment to the safety
program.
[[Page 535]]
(2) An effective institutional safety program requires a safety
officer appropriately trained in relevant safety technology. This
individual, besides supplying advice and recommendations, will ensure
that records are kept showing that the institution's physical facilities
and safety rules are internally consistent and compatible with potential
risks, as well as in compliance with all applicable laws, regulations,
and guidelines.
(3) The commander ensures safety in every department or other
equivalent administrative unit of the institution. Ensuring safe
operations is an integral function of each level of management through
the first line supervisor. The safety office staff must work closely
with administrators and investigators to develop and implement written
policies and practices that promote safe laboratory work. Collectively,
this group routinely must monitor current operations and practices, see
that appropriate audits are maintained, and continue to seek ways to
improve the safety program.
(4) Safety is a critical job element for each member of the
scientific and technical staff. Each individual working in the
laboratory must perform his or her job in a manner consistent with
safety policy and training.
(5) If laboratory goals dictate operations or substances not suited
to the existing facilities or equipment, the laboratory supervisor will,
assisted by the safety officer, advise and assist the laboratory worker
in developing or obtaining adequate facilities or equipment and
designing appropriate work procedures.
(6) The supervisor will authorize each specific operation, delineate
appropriate safety procedures, and instruct those who carry out the
operation.
(7) Potential hazards will be identified before work with etiologic
agents begins, and actions necessary to avoid accidents and illnesses
will be implemented. This practice, called a job safety analysis,
consists of breaking a job down into its logical steps, analyzing each
for its hazard potential, and deciding the safe procedures to use. The
process will be designed by a project director with input from
employees, and each step with potential for exposure or other incidents
must be described in writing in a standing operating procedure (SOP).
All such SOPs will be approved by, at a minimum, the commander or
institute director and the safety officer.
(8) The job safety analysis will include a consideration of health
hazards identified in AR 40-10 and of maximum credible events as
described in paragraph 2-8, AR 385-69.
(b) Safety plans. Clearly defined, published safety rules and
monitoring procedures for compliance must be established. These rules
will be readily available, in writing, for all involved in laboratory
operations. This goal may be accomplished by preparing or modifying a
facility safety plan, laboratory safety manual, occupational safety and
health program or equivalent. This plan will--
(1) Be coordinated with institutional and Federal, State, and local
emergency services.
(2) Be practiced with the emergency groups whose services are part
of that plan prior to any need for their services, so that they can
become familiar with any potential problem areas that may be encountered
when they are called upon for assistance.
(3) Describe the method of rapid communication (for telephone,
alarms, and so forth) that will be used during an emergency.
(4) Describe the institution's etiologic agent labeling system.
(5) Describe the institution's requirements for testing engineering
controls (for example, biological safety cabinets and high efficiency
particulate air (HEPA) filters) and essential safety equipment (for
example, autoclaves) that are used to conduct RDTE funded by the BDP.
(6) Appoint and train personnel responsible for handling an
emergency.
(7) Require that emergency telephone numbers be posted, so that
emergency service personnel know whom to contact at all times of the day
or night.
(8) Describe the institution's rules that have been established and
are practiced to limit access to the facilities where etiologic agents
under the sponsorship of the BDP are handled. The rules will include the
following requirements:
[[Page 536]]
(i) Access to biosafety level (BL)-1 and BL-1 large-scale (LS)
laboratories is limited or restricted at the discretion of the commander
or institute director when experiments are in progress.
(ii) Access to areas classified as BL-2, BL-2 LS, or where work with
toxins is conducted, is limited by the commander or institute director
when work with etiologic agents is in progress. Individuals who are at
increased risk of acquiring infection or for whom infection may be
unusually hazardous are not allowed in the laboratory. Only persons who
have been advised of the potential hazard and meet any specific entry
requirements (for example, immunization) may enter the individual
laboratory or animal rooms. The commander or institute director must
assess each circumstance and determine who may enter or work in the
laboratory.
(iii) Access to areas classified as BL-3 or BL-3 LS is limited as
stated in Sec. 627.7(b)(8)(ii), and is restricted to those persons whose
presence in the facility or individual laboratory rooms is required for
program or support purposes. Individuals under 18 years of age may not
enter the controlled area.
(iv) Access to BL-4 facilities is limited as stated in
Sec. 627.7(b)(8) (ii) and (iii). This is done with secure, locked doors
with access controlled by the commander or institute director, safety
officer, or other person responsible for the physical security of the
facility. Before entry, all persons will be advised as to the
appropriate safeguards for ensuring their safety. Authorized persons
must comply with these instructions and all other applicable entry and
exit procedures. A logbook will be maintained for all personnel to
indicate the date and time of each entry and exit. A card-key activated
computer record (or other electronic entry device) may be used if it
indicates the date and time of both entry and exit.
(9) Describe the system that is developed and is operational for the
reporting of accidents and exposures, employee absenteeism, and for the
medical surveillance of potential laboratory-associated illnesses.
(c) Safety meetings and safety committees. In effective safety
programs, everyone associated with the laboratory becomes involved. This
is done by ensuring maximum participation in planning and by conducting
group safety meetings.
(1) A staff safety committee, consisting of the commander or
institute director or his or her designated representative, research
supervisors, managers, medical personnel, employees, and the safety
officer, will be established. This group leads the safety effort,
reviews mishaps, and recommends changes in policies, safety program, or
equipment as needed to improve safety.
(2) Safety committees will meet at least quarterly and minutes will
be prepared and maintained for at least 3 years.
(3) When work with recombinant DNA molecules is undertaken, an
institutional biosafety committee (IBC) for review of such work will be
established and will function as stated in the NIH Guidelines for
Research Involving Recombinant DNA Molecules (see appendix A to this
part).
(d) SOPs. Besides the documented safety program that will be in
effect, each institution will require that an SOP be established for
each unique biological defense RDTE operation. The SOPs will meet the
criteria stated in AR 385-69 and be reviewed and updated annually. A
copy of the SOP will be maintained in the work area. In addition, SOPs
will address the following issues--
(1) The unique hazards introduced by the activity in the work area.
(2) The methods of controlling these hazards.
(3) Any unique procedures and requirements needed that are not
described as universally required in the safety plan (for example,
signs, waste disposal, immunizations, emergency procedures, and
personnel monitoring).
(4) Specialized orientation or training of personnel beyond that
required in the safety plan.
(5) Ways of ensuring that the unique procedures are followed.
(6) Emergency procedures.
(e) Safety communications. Safety communications alert people to
newly recognized hazards, remind them of
[[Page 537]]
basic biological safety principles, and instill positive attitudes
toward safety. Training requirements are also found in Sec. 627.10(b). A
system of communication will be established to--
(1) Implement a biological safety training program for all personnel
working with hazardous biological or chemical materials.
(2) Publish information addressing useful biological safety advice
and accounts of laboratory accidents, along with the lessons to be
learned from them.
(3) Make reference books and regulations concerning laboratory
hazards, occupational health, and proper laboratory practices readily
available.
(4) Assure that material safety data sheets (MSDS) for hazardous
chemicals used in the laboratory are readily available to all employees.
(f) Safety audits. One of the essential elements of a good safety
program is the conduct of periodic audits of the safety performance in a
laboratory. Observing individual safety practices and checking the
operability of safety equipment and compliance with safety rules must be
part of the audit.
(1) An individual and an alternate will be appointed for each
laboratory or room where BDP work is conducted. On a daily basis he or
she will monitor the conduct of personnel within their room(s) and
maintenance of the room to see that they comply with the safety program
and SOPs.
(2) Supervisors will ensure that their projects comply with
applicable safety requirements and will audit their areas at least
weekly to ensure compliance.
(3) The safety officer or his or her qualified designee will inspect
the institution's BL-1, BL-2, and toxin laboratories quarterly. BL-3 and
BL-4 laboratories and those in which dry forms of highly potent toxins
are handled will be inspected monthly by safety and health
professionals. These inspections will be announced and include coverage
of general safety practices as well as features specific to a particular
biosafety level.
(i) Reports of deficiencies or procedures that create a potentially
life-threatening situation will be made directly to supervisory
personnel and the commander or institute director and actions will be
taken immediately to correct the situation. The operation will not
continue until every deficiency is corrected.
(ii) Reports of deficiencies for other than life-threatening
situations will be made as soon as possible to the appropriate
supervisor, with copies furnished to the commander or institute
director. If a problem is widespread, all affected personnel will be
notified.
(4) Supervisory personnel notified of safety deficiencies by the
safety officer will ensure that the people directly concerned are
contacted and that the deficiencies are remedied before operations are
resumed.
(5) Malfunctioning equipment must be reported to the appropriate
individuals, labeled to indicate that it should not be used, and
repaired promptly.
(6) As a minimum, the audits conducted by the safety officer or his
or her qualified designee will cover the items listed in appendix C to
this part.
(g) Documentation. Records, documenting the following items, will be
maintained for 3 years:
(1) Safety audits and the corrective measures.
(2) Risk assessments for proposed new laboratory procedures.
(3) Annual reviews of established SOPs.
(4) Training.
(5) Engineering controls and protective equipment certifications and
tests.
(6) Safety committee meeting minutes and recommendations.
(7) Any outside auditor comments and responses.
Sec. 627.8 Occupational health.
An occupational health program will be implemented per AR 40-5,
chapter 5, for all employees whose employment requires that they conduct
duties in a BDP etiologic agent area. Essential elements of the program
will include--
(a) Medical surveillance examinations. Medical examinations by a
licensed medical doctor will be given prior to employment, at least
every 3 years thereafter, and upon termination of duties requiring
access to laboratories where etiologic agents are used. When full
medical examinations are not given annually, health professionals will
perform annual health screening.
[[Page 538]]
Safety and health professionals will ensure that medical examiners are
made aware of all hazardous substances each employee works with at the
time of the medical examination. The physician's findings will include
assessment of whether an employee has any health condition that would
preclude work with etiologic agents. If any of the findings obtained
during the examination are outside the normal range, the employee's
supervisor and the employee will be notified and counseled on the
courses of action available. In addition, a safety and health audit will
be conducted to identify any potential occupational causes for the
abnormalities, and corrective measures will be taken if applicable.
(b) Serum samples. When appropriate, considering the agent(s)
handled, baseline serum samples for laboratory and other at-risk
personnel will be collected and stored for their biologically useful
lifetime, but not longer than 40 years. Additional serum specimens will
be collected periodically, based upon the agents handled, or as required
by participation in a special immunizations program. SOPs will be
written detailing the collection procedures and periods if serum
sampling is deemed necessary.
(c) Assignment of personnel. Personnel assigned duties in work areas
where etiologic agents are used will be evaluated to determine their
suitability for their assigned tasks by the installation medical
authority. Only personnel who are physically and mentally capable of
working in biocontainment areas (BL-3 and BL-4) or with toxins will be
assigned to these duties.
(d) Immunization of at-risk personnel. The guidelines for
immunizations in the latest edition of the American College of
Physicians' Guide for Adult Immunizations and recommendations of Health
and Human Services (HHS) in publication number (NIH) 88-8395 shall be
followed. A resource list for available immunizations for personnel at
risk is given in appendix B of this part.
(e) Reporting exposures. Spills and mishaps which result in
observable, known or potential exposures to etiologic agents will be
immediately reported to the supervisor, the safety officer, the
responsible medical personnel, and the commander. Appropriate medical
evaluation, surveillance, and treatment will be provided and written
records of these occurrences will be maintained for 40 years. A Med-16
report will be initiated (see AR 40-400).
(f) Quarantine. When etiologic agents designated as BL-4 by the CDC-
NIH in HHS publication no. (NIH) 88-8395, (or most recent edition) are
handled, a facility for the quarantine, isolation, and medical care of
personnel with potential or known laboratory-associated exposures will
be available.
Sec. 627.9 Medical records.
Army activities will maintain medical records in accordance with AR
40-66 and FPM 293-31 for all military and Department of the Army (DA)
civilian employees who work with etiologic agents under sponsorship of
the BDP.
Subpart C--Operational Requirements
Sec. 627.10 Personnel prerequisites.
(a) Medical. Before to assignment to work with etiologic agents,
personnel will be evaluated by the appropriate medical personnel with
respect to their assignment and will be evaluated in the medical
surveillance program described in Sec. 627.8.
(b) Training. All personnel directly or indirectly involved with
containment or handling of known and potentially biohazardous material
shall receive instruction that adequately prepares them for their
assigned duties. Training will be given by occupationally qualified
personnel as determined by the commander. This training will be
documented and will include--
(1) General training--
(i) Personal hygiene related to laboratory work.
(ii) Laboratory practices.
(iii) Personal protective equipment.
(iv) Effective use of engineering controls.
(v) Packaging, transportation, and shipment of etiologic agents
(when applicable).
(vi) Hazardous and infectious waste disposal, handling, and
minimization procedures.
[[Page 539]]
(2) Training conducted specifically for the facilities that the
individual will be working in, including--
(i) Procedures for the facility.
(ii) Reporting incidents and accidents.
(iii) Labeling and posting of signs.
(iv) Biohazardous waste handling, approaches to minimizing the
volume of waste, decontamination, packaging, and disposal.
(v) Emergency procedures.
(3) Additional general training required for work in facilities
where viable etiologic agents are present.
(i) Aseptic technique and procedures to include hands-on instruction
and demonstration of proficiency.
(ii) Concept and definition of biosafety levels.
(iii) Disinfection and sterilization.
(iv) Safe use of workplace equipment, for example autoclave and
centrifuge.
(v) Monitoring and auditing requirements.
(vi) Precautions for handling blood, tissues, and body fluids (when
applicable).
(vii) The infectivity, pathogenicity, mode(s) of transmission, and
medical surveillance requirements of specific agents.
(viii) Training for all new employees will include a period of
supervised orientation in the facilities by a scientist or technician
with specific training in the procedures and properties of the etiologic
agents in use. During the training period, new laboratory personnel will
be under the constant supervision of appropriately trained personnel.
(ix) Personnel who are assigned tasks in BL-2, BL-3, or BL-4
facilities will also have specific training in handling pathogens.
(x) Personnel assigned duties in a BL-4 facility will also have
specific and thorough training in handling extremely hazardous
infectious agents, the primary and secondary containment functions of
standard and special practices, use of personal protective equipment,
containment equipment, and laboratory design characteristics.
(4) Additional general training for handling toxins will include
relevant items from Sec. 627.10 plus--
(i) The availability of reference material on the hazards and safe
handling of toxic substances.
(ii) The biological effects of the toxin(s) in use.
Sec. 627.11 Operational prerequisites.
(a) Evaluation of the risks. The risk assessment of laboratory
activities involving the use of etiologic agents is ultimately a
subjective process. Those risks associated with the agent, as well as
with any adjunct elements of the activity to be conducted, (chemicals,
radioisotopes, end-products, and so forth) must be considered in the
assessment. The appropriate biosafety level for work with a particular
agent or animal study depends on the virulence, pathogenicity,
biological stability, route of transmission, and communicability of the
agent; the nature of the laboratory; the procedures and manipulations to
be used; the quantity and concentration of the agent; and the
availability of effective vaccines or therapeutic measures.
(b) The characteristics of etiologic agents, primary laboratory
hazards of working with the agent, and recommended biosafety levels are
described by CDC-NIH (HHS publication No. (NIH) 88-8395), the
considerations for recombinant DNA molecules are described by NIH, and
those for oncogenic viruses are described by NCI-NIH (sources listed
below). The commander or institute director will assign work with given
etiologic agents to the appropriate biosafety level. A risk assessment
should take into account not only the NIH Guidelines for Research
Involving Recombinant DNA Molecules, but also potential hazards
associated with the organism and the product of the experimentation.
(1) When established guidelines exist, these will be followed. The
primary source guidelines are--
(i) HHS Publication No. (NIH) 88-8395, Biosafety in Microbiological
and Biomedical Laboratories, as amended, and updates published in
Morbidity and Mortality Weekly Report.
(ii) NIH Guidelines for Research Involving Recombinant DNA Molecules
(FR 51: 16958-16985 and updates).
[[Page 540]]
(iii) The publication by the American Committee on Arthropod-Borne
Viruses Subcommittee on Arbovirus Laboratory Safety (SALS) entitled
Laboratory Safety for Arboviruses and Certain Other Viruses of
Vertebrates in the American Journal of Tropical Medicine and Hygiene,
29(6), 1980, pp. 1359-1381.
(iv) The Department of Health and Human Services Publication No.
(NIH) 76-1165 by the National Cancer Institute (NCI) entitled Biological
Safety Manual for Research Involving Oncogenic Viruses.
(2) When samples with unidentified viable agents are obtained, a
knowledgeable and qualified scientist will evaluate the risks and make
recommendations to the safety officer, who will add recommendations for
review and approval by the commander or institute director. When
guidelines for a specific organism are not established, in addition to
these steps, the CDC or SALS or both will be consulted. Their
recommendations will be documented and provided to the commander or
institute director before approval.
(c) Selection of facilities. The facility requirements identified by
the risk assessment will be adhered to. Any variations and compensatory
measures will be approved by the IBC (when recombinant DNA molecules are
involved), the safety officer, and the commander or institute director
before a request for an exception or waiver is submitted as stated in AR
385-69.
(d) Policies and procedures. Policies in the form of a laboratory
safety manual, regulations, memorandums, or SOPs are required for work
with etiologic agents in the BDP. Before beginning a new procedure, the
policies and procedures will be reviewed to ascertain that the intended
operations are described and to determine the requirements that apply to
the operation. If procedures exist for the intended operation, personnel
will be trained to follow them; if procedures do not exist, then a
detailed SOP will be written, reviewed, and approved before beginning
the operation. SOPs will conform to the requirements stated in
Sec. 627.7(d), and be signed by all personnel who are required to follow
the procedures, thus acknowledging that they have read and understood
the contents. All SOPs that pertain to a specific area (room,
laboratory, or suite) will be available at the worksite.
Sec. 627.12 General laboratory techniques.
The general requirements for use of etiologic agents are composed of
two sets of requirements, with the requirements for toxins being a
subset of the requirements for handling viable etiologic agents. These
requirements are as follows--
(a) General techniques applicable to etiologic agents.
(1) A fully fastened long-sleeved laboratory coat, gown, uniform, or
coveralls will be worn in laboratories or animal rooms.
(2) Eating, drinking, smoking, and applying cosmetics are not
permitted in the work areas.
(3) Personnel must wash their hands after they handle etiologic
agents or animals, and before leaving the laboratory area.
(4) Mouth pipetting is strictly prohibited. Mechanical pipetting
aids must be used.
(5) Gloves--(i) Will be worn when manipulating etiologic agents and
handling containers of etiologic agents. Gloves are not required when
materials are packaged appropriately for shipment.
(ii) Will be selected based on the hazards.
(iii) Will be changed frequently (or decontaminated frequently), and
will be decontaminated or discarded into a labeled biohazard container
after each use and immediately upon observable direct contact with an
etiologic agent.
(iv) Will be removed at the work-space (workbench or hood) after
handling etiologic agents to ensure that doorknobs and other surfaces
are not contaminated.
(6) Good housekeeping will be maintained. This includes--
(i) Work areas free of clutter.
(ii) Work environment free of tripping hazards, with adequate access
to exits, emergency equipment, controls, and such.
(iii) Benches and general work areas will be cleaned regularly using
a wet
[[Page 541]]
sponge or similar method with disinfectant as appropriate. Methods that
stir up dust such as sweeping or using vacuum cleaners, (except for
HEPA-filtered vacuum cleaners) are unacceptable.
(iv) Specific work areas will be cleaned and decontaminated
immediately following each use of an etiologic agent (at least once a
day) and after any spill of viable material.
(v) Hallways and stairways will not be used for storage.
(7) All solutions, reagents, and chemicals will be labeled.
(8) All contaminated liquid or solid wastes will be inactivated
before disposal.
(9) Work will be conducted over spill trays or plastic-backed
absorbent paper. The paper will be removed, decontaminated, or
disinfected, and the general area wiped with decontaminant at the end of
each day or at the end of the experiment, whichever occurs first.
(10) Etiologic agents will be kept in closed containers when not in
use. Cultures, solutions, or dried etiologic agents in glass vessels
transported or incubated within a room or suite will be handled in
nonbreakable, leak-proof pans, trays, pails, carboys, or other secondary
containers large enough to contain all the material, if the glass vessel
leaks or breaks. Etiologic agents removed from a room or suite for
transport to another approved area within the same building will be
placed in a closed unbreakable secondary container before removal from
the laboratory. The secondary container will be labeled on the exterior
with a biohazard symbol and identification of the contents, including
the required biosafety level, the scientific name, the concentration (if
applicable), and the responsible individual. The secondary containers
will be wiped with suitable disinfectant before removal from the
laboratory or area.
(11) Working stocks of etiologic agents will be stored in double
containers. The primary and secondary containers will provide a positive
seal and the secondary container will be unbreakable. The secondary
container will be labeled as stated in Sec. 627.12 (a)(10) and with the
date stored.
(12) Storage units (for example, freezers, refrigerators, cabinets,
and hoods) will be labeled with the universal biohazard sign and
indicate the classes of etiologic agents contained in them. Storage
units will be secured when not in use.
(13) All contaminated materials, containers, spills, and solutions
will be decontaminated or disinfected by approved methods before
disposal.
(14) After injection of an etiologic agent into animals, the site of
injection will be swabbed with a decontaminant.
(15) Syringes. (i) Reusable or disposable syringes will be of the
fixed needle or LUER-LOK type (or equivalent) to assure that the needle
cannot separate during use.
(ii) After use, nondisposable glass syringes with attached needles
contaminated with etiologic agents will be submerged in a container of
decontaminant. Disposable syringes will be discarded with needles
attached in puncture-proof rigid containers. Needles will not be
recapped after use.
(iii) Sterilized or decontaminated containers marked ``Syringes and/
or Needles'' may be deposited in appropriate refuse containers after
proper packaging and destruction of the contents.
[Note: Many States, especially those on the Eastern seaboard, have
implemented strict requirements for the disposal of medical wastes. For
example, Maryland has designated all waste from a microbiological
laboratory as hazardous waste with licensing requirements for generators
of 50 kilograms per month or more of waste, while all medical waste
released for transport off-site must be manifested to a State licensed
medical waste hauler with the destination specified. Additionally, in
some cases, the local government (for example, a city) regulates the
disposal of these wastes. These requirements will be identified and
followed.]
Needles or syringes may not be destroyed by clipping. A mechanical shear
may be used to smash or sheer needles after or concurrently with
sterilization or decontamination.
(16) Refrigerators, deep freezers, and dry ice chests should be
checked, cleaned out, and defrosted periodically to remove any ampules,
tubes, and so forth, containing etiologic agents that may have broken
during storage. Rubber gloves and respiratory protection
[[Page 542]]
appropriate to the materials in storage should be worn during cleaning.
Do not store flammable solutions in nonexplosion proof refrigerators.
(b) Additional techniques applicable to work with viable etiologic
agents. The major objective of these techniques is to assist in
protection against laboratory acquired infections. Air sampling studies
have shown that aerosols are generated from most of the manipulations of
bacterial and viral cultures common to research laboratories. The
generation of aerosols during routine laboratory manipulations must be
considered when evaluating the individual degree of risk, keeping in
mind the four main factors governing infection: dosage, virulence of the
organism, route of infection (for example, skin, eyes, mouth, lungs),
and host susceptibility (for example, state of health, natural
resistance, previous infection, response to vaccines and toxoids). The
requirements stated below are minimum handling requirements to prevent
accidental infection created by incidental aerosols.
(1) All procedures are performed carefully to minimize the creation
of aerosols.
(2) No infectious mixtures will be prepared by bubbling air through
a liquid.
(3) Pipettes.
(i) No infectious material will be forcibly ejected from pipettes.
Only to deliver (TD) pipettes will be used.
(ii) Pipettes used with infectious or toxic materials will be
plugged with cotton unless they are used exclusively in a gas-tight
cabinet system.
(iii) Contaminated pipettes will be placed horizontally in a rigid
container containing enough disinfectant for complete immersion.
Cylinders used for vertical discard are not recommended. The container
and pipettes must be autoclaved as a unit and replaced by a clean
container containing fresh disinfectant.
(iv) Pipetting devices must be used. Under no circumstances is mouth
pipetting permitted.
(4) Syringes. (i) Using syringes and needles for making dilutions of
etiologic agents is not recommended.
(ii) When removing a syringe and needle from a rubber stopper bottle
containing viable etiologic agents, an alcohol soaked pledget around the
stopper and needle will be used.
(iii) Excess fluid and bubbles should be expelled from syringes
vertically into a cotton pledget soaked with disinfectant or into a
small bottle containing disinfectant-soaked cotton.
(iv) The site of injection of an animal will be swabbed with a
disinfectant before and after injection.
(v) After use, syringes contaminated with residual infectious fluid
will be submerged in a container of disinfectant in a safety cabinet
prior to removal for autoclaving. To minimize accidental injection of
infectious material, the removable needles should remain on such
syringes until after autoclaving. When possible, syringes with attached
needles should be placed in a pan separate from that holding other
discarded materials.
(vi) Caps will not be placed over needles until after disinfection.
During recapping, procedures to prevent personal injuries will be used.
(5) Centrifuges and shakers. (i) Before centrifuging, tubes, rotors,
seals, and gaskets will be checked for cleanliness and integrity. In low
speed clinical-type centrifuges, a germicidal solution may be added
between the tube and trunnion cup to disinfect the outer surfaces of
both and to cushion against shocks that might break the tube. Metal or
plastic tubes (other than nitro-cellulose) will be used.
(ii) Decanting from centrifuge tubes will be avoided. If decanting
is necessary, the outer rim will be wiped with a disinfectant after
decanting so that material on the lip cannot spin off as an aerosol.
Centrifuge tubes will not be filled byond the level the manufacturer
recommends.
(iii) Broth cultures will be shaken in a manner that avoids wetting
the plug or cap.
(6) Water baths in which viable etiologic agents are incubated must
contain a disinfectant. For cold water baths, 70 percent propylene
glycol is recommended. The disinfectant should be changed frequently.
(7) When a laboratory vacuum is used to manipulate viable etiologic
agents, a secondary reservoir containing disinfectant and a HEPA filter
must be employed to ensure that the laboratory
[[Page 543]]
vacuum lines do not become contaminated.
(8) Test tubes. (i) Tubes containing viable etiologic agents should
be manipulated with extreme care. Studies have shown that simple
procedures, such as removing a tube cap or transferring an inoculum, can
create a potentially hazardous aerosol.
(ii) Manipulation of biohazardous test tubes will be conducted in
biological safety cabinets. Tubes and racks of tubes containing
biohazardous material should be clearly marked. The individual employee
must ensure that tubes containing biohazardous material are properly
sterilized prior to disposal or glassware washing. Safety test tube
trays should be used in place of conventional test tube racks to
minimize spillage from broken tubes. When safety test tube trays are not
used, the conventional test tube racks will be placed in a tray large
enough to contain any potential spill. A safety test tube tray is one
having a solid bottom and sides deep enough to hold all liquids, should
a test tube break.
(9) Care should be exercised when using membrane filters to obtain
sterile filtrates of viable etiologic agents. Due to the fragility of
the membranes and other factors, such filtrates cannot be considered
noninfectious until laboratory culture or other tests have proven their
sterility.
(10) The preparation, handling, and use of dry powders of viable
etiologic agents in open containers presents unusual hazards. The
slightest manipulation of such powders can cause the generation of
aerosols containing a high concentration of etiologic agents. Therefore,
work with dry powders of etiologic agents in open containers should be
carried out in gas-tight biological safety cabinets.
Sec. 627.13 Biosafety level 1.
(a) Requirements beyond those for all etiologic agents. BL-1
operations follow the general techniques described in Secs. 627.12(a)
and 617.12(b).
(b) Additional laboratory requirement. Contaminated materials that
are to be decontaminated at a site away from the laboratory are placed
in a durable leak-proof container which is closed before being removed
from the laboratory. Examples of suitable containers are metal tubs with
lids or plastic bags that are sealed and then placed inside a rigid
container for transport.
(c) Additional animal requirements. (1) Bedding materials from
animal cages will be removed in such a manner as to minimize the
creation of aerosols and disposed of in compliance with applicable
institutional or local requirements.
(2) Cages are washed manually or in a cagewasher. Temperature of
final rinse water will be a minimum of 180 deg.F.
(3) Laboratory coats, gowns, or uniforms worn in animal rooms shall
not be worn in other areas.
Sec. 627.14 Biosafety level 2.
(a) Additional requirements. In addition to the general
microbiological techniques stated in Sec. 627.13, BL-2 operations
include the following requirements:
(1) When etiologic agents are in use, a hazard warning sign
incorporating the universal biohazard symbol is posted on the access
door of the work area. The hazard warning sign identifies the etiologic
agent, lists the name and telephone number of the institute director or
other responsible person(s), and indicates the special requirement(s)
for entering the laboratory.
(2) Animals not involved in the work being performed are not
permitted in the laboratory.
(3) Special care is taken to avoid skin contamination with the
etiologic agents; gloves will be worn when handling etiologic agents or
infected animals.
(4) All wastes from laboratories and animal rooms are decontaminated
before disposal.
(5) Hypodermic needles and syringes are used only for parenteral
injection and aspiration of fluids from laboratory animals and diaphragm
bottles.
(6) Spills and accidents which result in a potential exposure to
etiologic agents will be reported immediately to the safety officer, the
project leader, and the institute director.
(7) Biological safety cabinets (Class I or II) will be used when:
(i) Procedures with a high potential for creating infectious
aerosols are conducted.
[[Page 544]]
(ii) High concentrations or large volumes of etiologic agents are
used.
(8) Laboratory coats, gowns, smocks, or uniforms will be removed
before leaving the animal facility or laboratory area.
(b) Additional animal requirements.
(1) Cages must be decontaminated, preferably by autoclaving, before
they are cleaned and washed.
(2) Approved molded masks are worn by all personnel entering animal
rooms housing nonhuman primates.
(3) If floor drains are provided, the drain traps will be kept
filled with water or a suitable disinfectant.
Sec. 627.15 Biosafety level 3.
(a) Additional requirements. In addition to the requirements stated
in Secs. 627.13 and 627.14, the following requirements apply--
(1) Approved molded masks or respirators with HEPA filters are worn
by all personnel in rooms housing infected animals.
(2) Protective clothing worn in a laboratory or animal room will be
removed before exiting the laboratory or animal room.
(3) Clothing worn in laboratories and animal areas to protect street
clothing will be decontaminated before being laundered.
(b) Additional laboratory requirements. (1) Laboratory doors will be
kept closed.
(2) All activities involving etiologic agents will be conducted in
biological safety cabinets (Class I, II, or III) or other physical
containment devices within the containment module. No work in open
vessels is conducted outside a biological safety cabinet.
(3) The work surfaces of biological safety cabinets and other
containment equipment will be decontaminated after work with etiologic
agents. Plastic-backed paper toweling should be used on nonperforated
work surfaces within biological safety cabinets to facilitate clean-up.
(c) Additional animal requirements. (1) Cages are autoclaved before
bedding is removed and before they are cleaned and washed.
(2) Gloves are removed aseptically and autoclaved with other wastes
before being disposed of or reused.
(3) Boots, shoe covers, or other protective footwear and
disinfectant foot baths must be available and used when indicated.
(4) Personal protective clothing and equipment and other physical
containment devices are used for all procedures and manipulations of
etiologic agents or infected animals. The risk of infectious aerosols
from infected animals or their bedding shall be reduced by housing
animals in partial containment caging systems as described in
Sec. 627.56.
(d) Work with BL-3 etiologic agents that require additional
secondary containment. Facilities in which work with certain viruses,
for example, Rift Valley fever, yellow fever, and Venezuelan equine
encephalitis, is conducted require HEPA filtration of Xallexhaust air
prior to discharge from the laboratory. All persons working with those
agents for which a vaccine is available should be immunized.
Sec. 627.16 Biosafety level 4.
Laboratory work at BL-4 must follow the requirements stated in
Secs. 627.13, 627.14 and 627.15 as well as the following:
(a) All activities are conducted in Class III biological safety
cabinets or in Class I or II biological safety cabinets in conjunction
with a one-piece positive pressure personnel suit ventilated by a life-
support system.
(b) Biological materials to be removed from the Class III cabinet or
from the maximum containment laboratory in a viable or intact state must
be transferred to a sealed nonbreakable primary container, enclosed in a
nonbreakable sealed secondary container, and removed from the facility
through a disinfectant dunk tank, fumigation chamber, or an airlock
designed for this purpose.
(c) No materials, except for biological materials that are to remain
in a viable or intact state, are removed from the maximum containment
laboratory unless they have been autoclaved or decontaminated before
they leave the facility. Equipment or material which might be damaged by
[[Page 545]]
high temperature or steam is decontaminated by gaseous or vapor methods
in an airlock or chamber designed for this purpose.
(d) Personnel may enter and leave the facility only through the
clothing change and shower rooms. Personnel must shower each time they
leave the facility. Personnel may use the airlocks to enter or leave the
laboratory only in an emergency.
(e) Street clothing must be removed in the outer clothing change
room and kept there. Complete laboratory clothing, including
undergarments, pants and shirts or jumpsuits, shoes, and gloves, will be
provided and must be used by all personnel entering the facility. Head
covers are provided for personnel who do not wash their hair during the
shower. When leaving the laboratory and before proceeding into the
shower area, personnel must remove their laboratory clothing and store
it in a locker or hamper in the inner change room.
(f) When etiologic agents or infected animals are present in the
laboratory or animal rooms, a hazard warning sign incorporating the
universal biohazard symbol must be posted on all access doors. The sign
must identify the agent, list the name of the commander or institute
director or other responsible person(s), and indicate any special
requirements for entering the area (for example, the need for
immunizations or respirators).
(g) Supplies and materials needed in the facility are brought in by
way of the double-doored autoclave, fumigation chamber, or airlock which
is appropriately decontaminated after each use. After securing the outer
doors, personnel within the facility retrieve materials by opening the
interior doors of the autoclave, fumigation chamber, or airlock. These
doors are secured after materials are brought into the facility.
(h) Materials (for example, animals and clothing) not related to the
experiment being conducted are not permitted in the facility.
(i) Whenever possible, avoid using any glass items.
Sec. 627.17 Toxins.
The laboratory facilities, equipment, and procedures appropriate for
work with toxins of biological origin must reflect the intrinsic level
of hazard posed by a particular toxin as well as the potential risks
inherent in the operations performed. All toxins must be considered to
pose a hazard in an aerosol form. However, most toxins exert their
effects only after parenteral exposure or ingestion, and a few toxins
present a dermal hazard. In general, toxins of biological origin are not
intrinsically volatile. Thus, the laboratory safety precautions
appropriate for handling these materials closely parallel those for
handling infectious organisms. The requirements in this section for the
laboratory use of toxins of biological origin include the requirements
in Sec. 627.12(a) and the following:
(a) Vacuum lines. When vacuum lines are used with systems containing
toxins, they will be protected with a HEPA filter to prevent entry of
toxins into the lines (or sink drains when water aspirators are used).
(b) Preparation of concentrated stock solutions and handling closed
primary containers of dry toxins. Preparation of primary containers of
toxin stock solutions and manipulations of closed primary containers of
dry forms of toxins will be conducted--
(1) In a chemical fume hood, a glove box, or a biological safety
cabinet or equivalent containment system approved by the safety officer.
(2) While wearing eye protection if using an open-fronted
containment system.
(3) Ensuring that gloves worn when handling toxins will be disposed
of as toxin waste, with decontamination if required.
(4) With the room door closed and posted with a universal biohazard
sign, or other sign, indicating that toxin work is in progress.
Extraneous personnel shall not be permitted in the room during
operations.
(5) Ensuring that toxins removed from hoods or biological safety
cabinets are double-contained during transport.
(6) After verification of hood or biological safety cabinet inward
airflow is
[[Page 546]]
made by the user before initiating work.
(7) Within the operationally effective zone of the hood or
biological safety cabinet.
(8) Ensuring that nondisposable laboratory clothing is
decontaminated before release for laundering.
(9) Ensuring that all individuals who handle toxins wash their hands
upon each exit from the laboratory.
(10) With two knowledgeable individuals present whenever more than
an estimated human lethal dose is handled in a syringe with a needle.
Each must be familiar with the applicable procedures, maintain visual
contact with the other, and be ready to assist in the event of an
accident.
(c) Manipulations with open containers of dry forms of toxins.
Handling dry forms of toxins in uncovered containers (for example,
during weighing) will be performed following the requirements stated in
Secs. 627.12(a), 627.17 (a) and (b), and the following:
(1) Manipulations will be conducted in a HEPA filtered chemical fume
hood, glove box, or biological safety cabinet. In addition the exhaust
may be charcoal filtered if the material is volatile.
(2) When using an open-fronted fume hood or biological safety
cabinet, protective clothing, including gloves and a disposable long-
sleeved body covering (gown, laboratory coat, smock, coverall, or
similar garment) will be worn so that hands and arms are completely
covered. Eye and approved respiratory protection is also required. The
protective clothing will not be worn outside of the laboratory and will
be disposed of as solid toxin waste.
(3) Before containers are removed from the hood, cabinet, or glove
box, the exterior of the closed primary container will be decontaminated
and placed in a clean secondary container.
(4) When toxins are in use, the room will be posted to indicate
``Toxins in Use--Authorized Personnel Only.'' Any special entry
requirements will be posted on the entrance(s) to the room.
(5) All operations will be conducted with two knowledgeable
individuals present. Each must be familiar with the applicable
procedures, maintain visual contact with the other, and be ready to
assist in the event of an accident.
(6) Individuals handling toxins will wash their hands upon leaving
the laboratory.
(d) Additional considerations of specific toxin properties. The
following requirements are in addition to the requirements stated in the
paragraphs above. Determine whether the material fits Sec. 627.17 (b) or
(c), and complies with the appropriate section and the following when
applicable:
(1) When handling dry forms of toxins that are electrostatic--
(i) Do not wear gloves (such as latex) that help to generate static
electricity.
(ii) Use glove bag within a hood or biological safety cabinet, a
glove box, or a class III biological safety cabinet.
(2) When handling toxins that are percutaneous hazards (irritants,
necrotic to tissue, or extremely toxic from dermal exposure)--
(i) Gloves will be selected that are known to be impervious to the
toxin and the diluent (when applicable) for the duration of the
manipulations.
(ii) Disposable laboratory clothing will be worn, left in the
laboratory upon exit, and disposed of as solid toxin waste.
(e) Aerosol exposures. The requirements found in Sec. 627.17 (a) and
(b) will be complied with plus the following:
(1) Chambers, nose-only exposure apparatus, and generation system
must be placed inside a fume hood, glove box, or a Class III biological
safety cabinet. Glove boxes and Class III biological safety cabinets
will have HEPA filters on both inlet and outlet air ports.
(2) The atmosphere from within the exposure chamber will be HEPA
filtered before release inside the hood, glove box, or cabinet.
(3) All items inside the hood, glove box, or Class III biological
safety cabinet will be decontaminated upon removal. Materials such as
experimental samples that cannot be decontaminated directly will be
placed in a closed secondary container, the exterior of which will be
decontaminated and labeled appropriately. Animals will have any areas
exposed to toxin wiped clean after removal from the exposure apparatus.
[[Page 547]]
(4) The interior of the hood, glove box, or cabinet containing the
chamber and all items will be decontaminated periodically, for example,
at the end of a series of related experiments. Until decontamintated,
the hood, box, or cabinet will be posted to indicate that toxins are in
use, and access to the equipment and apparatus restricted to necessary,
authorized personnel.
Sec. 627.18 Emergencies.
(a) Introduction. All laboratories will establish specific emergency
plans for their facilities. Plans will include liaison through proper
channels with local emergency groups and with community officials. These
plans will include both the building and the individual laboratories.
For the building, the plan must describe evacuation routes, facilities
for medical treatment, and procedures for reporting accidents and
emergencies. The plans will be reinforced by drills. Emergency groups
and community officials must be informed of emergency plans in advance
of any call for assistance. See AR 385-69.
(b) General emergency procedures. The following emergency procedures
will be followed for laboratory accidents or incidents--
(1) Using appropriate personal protection, assist persons involved,
remove contaminated clothing if necessary, decontaminate affected areas,
and remove personnel from exposure to further injury if necessary; do
not move an injured person not in danger of further harm. Render
immediate first aid if necessary.
(2) Warn personnel in adjacent areas of any potential hazards to
their safety.
(3) In case of fire or explosion, call the fire department or
community fire brigade immediately. Follow local rules for dealing with
incipient fire. Portable fire extinguishers will be made available with
instructions for their use. Fire fighters responding to the fire scene
will be advised to wear a self-contained positive pressure breathing
appartus to protect themselves from toxic combustion by-products.
(4) Laboratories must be prepared for problems resulting from severe
weather or loss of a utility service. In the event of the latter, most
ventilation systems not supplied with emergency power will become
inoperative. All potentially hazardous laboratory work must stop until
service has been restored and appropriate action has been taken to
prevent personnel exposure to etiologic agents.
(5) In a medical emergency, summon medical help immediately.
Laboratories without a medical staff must have personnel trained in
first aid available during working hours.
(6) For small-scale laboratory accidents, secure the laboratory,
leave the area, and call for assistance.
(7) When handling mixed hazards (for example, a substance or mixture
that may be infectious and radioactive, or infectious and chemically
toxic), respond with procedures addressing the greater hazard first, and
then follow through with those for the lesser hazards to ensure that all
appropriate steps have been taken.
(c) Evacuation procedures. Building and laboratory evacuation
procedures will be established and communicated to all personnel.
(1) Emergency alarm system. (i) There will be a system to alert
personnel of an emergency that requires evacuation of the laboratory or
building. Laboratory personnel must be familiar with the location and
operation of alarm equipment.
(ii) Isolated areas (for example, cold, warm, or sterile rooms) will
be equipped with an alarm or communication system that can be used to
alert others outside to the presence of a worker inside, or to warn
workers inside of an emergency that requires evacuation.
(2) Evacuation routes will be established and an outside assembly
area for evacuated personnel must be designated. All individuals should
be accounted for.
(3) Shut-down and start-up procedures.
(i) Guidelines for shutting down operations during an emergency
evacuation will be available in writing. Those guidelines will include
procedures for handling any power failure emergency.
(ii) Written procedures will also be provided to ensure that
personnel do not return to the laboratory until the emergency is ended.
Those procedures
[[Page 548]]
must also contain start-up operations for the laboratory.
(iii) All shut-down and start-up procedures will be available to
personnel and reviewed semiannually.
(4) All aspects of the building evacuation procedur will be tested
semiannually with practice drills.
(d) Spills. (1) All areas where work with etiologic agents is
performed will have designated personnel to respond to a spill and
provide protective apparel, safety equipment, and materials necessary to
contain and clean up the spill. Protective clothing requirements are
described in Sec. 627.21. Also, there will be supplies on hand to deal
with the spill consistent with the hazard and quantities of the spilled
substance.
(2) The safety officer will be notified immediately of all spills.
The first line supervisor will ensure that proper clean-up techniques
are employed.
(3) Etiologic agents. (i) A program for responding to spills of
etiologic agents will be developed and implemented. This program will
contain emergency response procedures for a biological spill, which will
be tailored to the potential hazard of the material being used, the
associated laboratory reagents involved, the volume of material, and the
location of the materials within the laboratory. Generally, the spill
should be confined to a small area while minimizing the substance's
conversion to an aerosol. The spill will be chemically decontaminated or
neutralized, followed by a cleanup with careful disposal of the residue.
If the spilled material is volatile and noninfectious, it may be allowed
to evaporate but must be exhausted by a chemical hood or ventilation
system.
(ii) When a mishap occurs that may generate an aerosol of etiologic
agents requiring BL-2 (or higher) containment, the room must be
evacuated immediately, the doors closed, and all clothing
decontaminated, unless the spill occurs in a class II or class III
biological safety cabinet. Sufficient time must be allowed for the
droplets to settle and the aerosols to be reduced by the air changes of
the ventilation system before decontaminating the area. The area will
then be decontaminated to prevent exposure to the infectious agents or
toxic substances. Reentry procedures to perform the decontamination will
conform to Sec. 627.18(e).
(iii) A spill of biohazardous material within a biological safety
cabinet requires a special response and cleanup procedure. Cleanup will
be initiated while the cabinet continues to operate, using an effective
chemical decontaminating agent. Aerosol generation during
decontamination and the escape of contaminants from the cabinet must be
prevented. Caution must be exercised in choosing the decontaminant,
keeping in mind that fumes from flammable organic solvents, such as
alcohol, can reach dangerous concentrations within a biological safety
cabinet.
(4) Combined radioactive and biological spills. (i) Both the
radiation protection officer (RPO) and the safety officer must be
notified immediately whenever there is a spill of radioactive biological
material, regardless of its size. Laboratory personnel may be expected
to clean up the spill. The RPO will direct the cleanup, in accordance
with the NRC license for the facility.
(ii) The spill will be cleaned up in a way that minimizes the
generation of aerosols and spread of contamination. All items used in
cleaning up the spill must be disposed of as radioactive waste.
(iii) Following cleanup, the area, affected protective clothing, and
all affected equipment and supplies must be surveyed for residual
radioactive contamination. All potentially affected areas and items that
are not disposable will be wipe-tested to verify that unfixed
radioactive contamination has been removed. If fixed contamination is
found, the RPO will determine the requirements for additional cleanup.
(e) Reentry procedures. This section applies when reentry is
necessary to clean up a spill outside of a hood or biological safety
cabinet, or to decontaminate or service engineering controls that have
failed or malfunctioned so that they do not provide the required
containment.
(1) When agents requiring BL-1 or BL-1 LS containment are involved,
the clothing requirements stated in Sec. 627.30
[[Page 549]]
(a) or (b) as appropriate will be followed. Individuals will remove the
required protective clothing when finished and wash their hands before
proceeding to other tasks.
(2) When agents requiring BL-2, BL-2 LS, or toxin procedures and
containment are involved, personnel will be required to wear the
clothing described in Sec. 627.30 (c) or (d) as appropriate. Outer
protective clothing will be removed and left in the room before exiting
and personnel will wash their hands before proceeding on to other
activities.
(3) When agents requiring BL-3, or BL-3 LS containment are involved,
containers for sealing up inner protective clothing and decontaminant
will be placed at the room exit. Personnel will be required to wear the
clothing described in paragraph 4-10e. When exiting the area after
decontamination procedures, individuals will remove their outer layer of
protective clothing just before exiting the room. Once outside the room,
the inner layer of protective clothing (for example, coverall) will be
removed and placed in the container and the inner gloves will be
decontaminated before being removed and placed in the container.
Personnel will proceed directly to the shower facility to take a
complete shower before exiting the facility.
(4) When agents requiring BL-4 containment are involved, the
following applies as appropriate to the type of BL-4 facility:
(i) When a spill requiring clean-up is in an area designed for use
with personal positive pressure suits, the entry and exit procedures
will be those normally required to enter or exit the area.
(ii) When entering a nonsuit area where a spill of etiologic agent
has occurred outside the containment of a Class III biological safety
cabinet, personnel will wear the clothing as described in
Sec. 627.30(f). Before entry, decontamination areas will be established.
To accomplish this, two step-in decontamination pans with the
appropriate disinfectant will be set up [one just inside the room (where
the contamination exists) and the second immediately outside the room].
Immediately outside the room, there will also be a sealable container
suitable for sealing up the suit and any air lines (if used).
(iii) When exiting the room, suited individuals will place all
equipment and other items in autoclaves or disinfectant, step into the
disinfectant pan, and wash down the exterior of their suits with
appropriate disinfectant. When completed, the door to the room will be
opened and the individual will step through the doorway into the second
disinfectant pan. The suit will be thoroughly rinsed with disinfectant
again before moving toward the exit from the facility. The suit (but not
the respirator) will be placed in the provided container. The individual
will proceed through another doorway before removing the respirator and
placing it in a closed container for decontamination. The individual
will then proceed directly to the shower area and take a full shower
before exiting the area. In case they are needed, personnel will be
standing by ready to render assistance. Suited individuals will be
visually observed, if possible. When visual observation is not possible,
a communications system is required.
(f) Mishap reports and investigations. (1) Each institution must
have a defined system for reporting laboratory injuries, illnesses, and
mishaps, as well as for investigating them. These events will be
documented and reported to the appropriate safety, supervisory, and
occupational health personnel. Those organizations subject to the
regulations promulgated by the OSHA will follow the specific
requirements for reporting injuries in the work place contained in those
regulations. The requirements stated in AR 385-69, State, and local
government requirements for similar reporting will be followed.
(2) Form(s) for recording mishaps will be available and completed
for all laboratory mishaps. Those reports must include a description of
the mishap and any factors contributing to it. In addition, a
description of any first aid or other health care given to the employee
will be included. Responsibility for completing these forms must be
clearly defined in the facility safety
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manual. Mishaps will be reviewed periodically by the safety officer, the
safety committee, the employee health unit, or other appropriate
personnel. Individual reports or a summary must be sent, along with
recommended changes in laboratory procedure or policy, to the commander
or institute director. Policy or procedural changes must be implemented
if deemed necessary by the commander or institute director.
(3) Any mishaps with etiologic agents used under sponsorship of the
BDP that result in sero-conversion or a laboratory-acquired illness will
be reported.
Sec. 627.19 Large-scale operations.
(a) Large-scale. In addition to the requirements stated in
Sec. 627.13, the following applies to research or production activities
involving viable etiologic agents in quantities greater than 10 liters:
(1) All large-scale operations will be conducted in facilities
described in Sec. 627.47.
(2) Cultures will be handled in a closed system.
(3) Sample collection, the addition of materials, and the transfer
of culture fluids shall be done in a manner which minimizes the release
of aerosols or contamination of exposed surfaces.
(4) A closed system or other primary containment equipment that has
contained viable organisms shall not be opened for maintenance or other
purposes unless it has been sterilized.
(5) SOPs will include a section describing and requiring a
validation of the process equipment's proper function.
(6) Scientists, technicians, equipment workers, and support
personnel with access to the large-scale production area during its
operation will be included in the medical surveillance program.
(b) BL-2--LS. In addition to the requirements stated in
Secs. 627.19(a) and 627.14, the following procedures will be employed
for BL-2--LS:
(1) Rotating seals and other mechanical devices directly associated
with the closed system used for the propagation and growth of viable
organisms shall be designed to prevent leakage or shall be fully
enclosed in ventilated housings that are exhausted through filters which
have efficiencies equivalent to HEPA filters or through other equivalent
treatment devices.
(2) A closed system used for the propagation and growth of viable
organisms and other primary containment equipment used to contain
operations involving viable organisms shall include monitoring or
sensing devices that monitor the integrity of containment during
operations.
(3) Systems used to propagate and grow viable organisms shall be
permanently identified. This identification shall be used in all records
reflecting testing, operation, and maintenance and in all documentation
relating to the use of this equipment.
(c) BL-3--LS. In addition to the requirements stated in
Secs. 627.19(a) and 617.14, the following procedures apply:
(1) Personnel entry into the controlled area shall be through the
entry area specified in Sec. 627.47(c)(1).
(2) Persons entering the controlled area shall exchange or cover
their personal clothing with work garments such as jumpsuits, long
sleeved laboratory coats, pants and shirts, head cover, and shoes or
shoe covers. On exit from the controlled area, the work clothing may be
stored in a locker separate from that used for personal clothing, or
discarded for laundering. Clothing shall be decontaminated before
laundering.
(3) Entry into the controlled area during periods when work is in
progress shall be restricted to those persons required to meet program
support needs.
(4) Prior to entry, all persons shall be informed of the operating
practices, emergency procedures, and the nature of the work conducted.
(5) The universal biohazard sign shall be posted on entry doors to
the controlled area and all internal doors. The sign posted on the entry
doors to the controlled area shall include a statement of agents in use
and personnel authorized to enter.
(6) Equipment and materials required for the management of accidents
involving viable organisms shall be available in the controlled area.
(d) BL-4--LS. Guidelines for these operations are not established.
If these
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are needed, they must be established by the United States Army Surgeon
General or the NIH on an individual basis.
Sec. 627.20 Operations with radioactive material.
Operations that combine etiologic agents with radioactive material
present unique problems. When this is the case, the following apply:
(a) Radiation program. A radiation program meeting the requirements
of AR 385-11 and NRC licensing that allows the particular isotope and
its use are required. The requirements for acquisition, handling
procedures, labeling, storage, training, monitoring, and disposal will
be described in an organization policy document.
(b) Procedure approval. In addition to the required approvals for
work with etiologic agents, the RPO will approve all SOPs involving the
use of radioactive materials. Laboratory operators must be fully
trained, with annual training updates as required by the existing
license.
(c) Special situations. (1) The laboratory waste must be segregated
as radioactive waste and disposed of as such after it has been
decontaminated. Do not mix nonradioactive waste with radioactive waste
as the disposal of radioactive waste is much more complex and expensive.
When RCRA-listed chemicals are mixed with radioactive waste, it becomes
``mixed waste'' for which there is currently no means of disposal.
(2) Activities conducted with radioisotopes should be confined to
the smallest number of areas or rooms consistent with requirements.
(3) Decontamination methods specific to etiologic agents will not
always remove radioactivity. Other methods, such as specialized
detergents and solvents designed for this use, should be employed to
remove residual radioactivity.
Subpart D--Personal Protective Equipment
Sec. 627.21 Introduction.
Personal protective equipment (PPE) includes clothing and equipment
used to protect the laboratory worker from contact with infectious,
toxic, and corrosive agents, as well as excessive heat, fire, and other
physical hazards. The appropriate PPE for any activity depends upon the
proposed operations and the potential hazards associated with them.
While PPE is an important item of personal protection, it serves as only
a secondary line of protection against hazards in the workplace.
Engineering controls (subpart H), combined with common sense, good
laboratory techniques, and adherence to SOPs, are the primary barriers
to exposure. There are some situations, however, in which it is either
impractical or impossible to rely exclusively on engineering controls.
In these cases, PPE may form the primary barrier between personnel and
the hazardous or infectious materials.
Sec. 627.22 Minimum laboratory attire for use of etiologic agents.
Individuals required to wear PPE will be trained in its proper use.
The PPE listed below is the minimum required when etiologic agents are
handled at any biosafety level. Research with etiologic agents usually
involves hazards other than those presented by the agents themselves.
When PPE is selected, the hazards presented by these other factors must
be considered regardless of the biosafety level used. For example, toxic
chemicals are commonly used in research involving etiologic agents. The
processes may expose personnel to physical hazards, such as heat or
animal bites, and the decontamination process may involve the handling
of toxic or corrosive materials. When the PPE required to mitigate these
hazards exceeds that of the minimum requirements, the necessary PPE will
be selected considering all the hazards. Information regarding the
additional appropriate PPE worn to protect against these hazards will be
available from one of the following sources: MSDS, SOP for the
operation, or the safety officer. Deviations from the standards stated
in approved SOPs must be approved by the safety officer. All laboratory
coats worn to protect the individual should be left in the laboratory
when that individual leaves. In each case, the minimum attire will be--
[[Page 552]]
(a) Laboratory workers. Street attire is permissible in the
laboratory, but must include closed-toe shoes. A full-length, long
sleeved, fully fastened laboratory coat, gown, or smock will be worn
over the street attire in the laboratory at all times. The laboratory
clothing will be removed and left in the laboratory when leaving to
enter nonlaboratory use areas.
(b) Animal caretakers. In addition to the clothing requirements in
Sec. 627.22(a), animal handlers will be provided with safety shoes or
safety boots. The requirements of Sec. 627.22(b) should also apply.
(c) Nonhuman primate rooms. Personnel entering rooms housing
nonhuman primates will wear the clothing stated in Sec. 627.22(a) and,
if applicable, Sec. 627.22(b) in addition to a molded mask or HEPA
filtered respirator, latex or vinyl gloves, and eye protection.
Sec. 627.23 Biosafety level 1.
This level requires only the minimum attire described in
Sec. 626.22.
Sec. 627.24 Biosafety level 2.
This level requires the following additions to the minimum clothing
specified in Sec. 627.22:
(a) Laboratory. Gloves (type dependent on the application) will be
worn when handling etiologic agents or containers of etiologic agents
and when handling infected animals.
(b) Animal rooms. (1) Protective clothing will be changed completely
every day. One- or two-piece laboratory suits or solid-front gowns and
wrap-around smocks are preferable. Full-length, long-sleeved, fully
fastened laboratory coats are allowed.
(2) Eye protection must be worn when handling nonhuman primates.
(3) Appropriate gloves must be worn.
(4) Molded masks or HEPA filtered respirators will be worn in rooms
housing nonhuman primates.
Sec. 627.25 Biosafety level 3.
The outer clothing worn in these facilities must never be worn
outside the facility. Color-coded clothing that is worn only in the
facility is recommended to remind individuals not to wear it outside.
The minimum clothing includes--
(a) Laboratory. (1) Long-sleeved, solid front, or wraparound gowns,
scrub suits, or coveralls over street attire which includes closed-toe
shoes. Dedicated shoes, boots, or shoe covers will be worn in the
facility.
(2) Appropriate gloves.
(b) Animal rooms. (1) A complete change of protective clothing on a
daily basis. Long-sleeved one- or two-piece solid front uniforms, solid-
front gown, wrap-around smocks, or solid front coveralls.
(2) Eye protection must be worn when handling nonhuman primates.
(3) Molded masks or HEPA filtered respirators will be worn in rooms
housing infected animals.
(4) Shoe covers will be worn and removed before exiting the room;
alternatively, disinfectant footbaths will be used for each exit from
the room when infected animals are present.
Sec. 627.26 Biosafety level 4.
Street clothing must be removed in an outer clothing change room and
kept there. Clothing worn in the facility will be removed in an inner
change room and a shower taken before replacing the street clothing. Two
distinct PPE requirements exist for BL-4 operations:
(a) Class III biological safety cabinet containment. Clothing
requirements when all etiologic agents and infected animals are housed
and manipulated in Class III biological safety cabinets will include--
(1) Complete change of clothing and wet shower upon exit. This
includes undergarments, pants and shirts or jump-suits, and shoes. While
it is preferred that the shower include washing the hair, head covers
will be worn by those who do not wash their hair on each exit.
(2) Appropriate inner gloves. The inner gloves will be donned in the
change room.
(b) Class I or II biological safety cabinet containment. Clothing
requirements for this level when etiologic agents are contained in Class
I or II biological safety cabinets of equivalent partial-
[[Page 553]]
containment caging systems (for infected animals) (See Secs. 627.56 and
627.57) include--
(1) Complete change of clothing and wet shower upon exit. This
includes undergarments, pants and shirts or jump-suits, and shoes. While
the shower should include washing the hair, head covers will be worn by
those who do not wash their hair on each exit.
(2) Appropriate inner gloves will be donned in the change room.
(3) A one-piece positive pressure suit described in Sec. 627.31(g).
(4) Impervious boots fitted over the suit.
Sec. 627.27 Large-scale (LS) operations.
The clothing requirements for these are the same as for the
corresponding biosafety levels for laboratory operations.
Sec. 627.28 Solutions of toxins and dry forms of toxins in closed containers.
In addition to the minimum clothing specified in Sec. 627.22,
disposable gloves or gloves designed to protect against the diluent will
be worn when handling these materials.
Sec. 627.29 Dry forms of toxins handled in open containers.
In addition to the requirements stated in Sec. 627.28, the
requirements stated in Sec. 627.18(c) apply.
Sec. 627.30 Situations specified in Sec. 627.18(e).
The clothing requirements for this section are for the emergency
procedures specified in Sec. 627.18(e). Because situations can occur and
there is no feasible or available means to mitigate the potential hazard
adequately by engineering controls, the clothing requirements exceed
those required for a properly conducted laboratory operation at an
equivalent biosafety level. The protective equipment required will be
selected based upon an assessment of the potential hazards that could be
encountered. The following clothing requirements are given as a guide.
The selection of PPE will be based upon the highest possible level of
contamination that could exist in the room. This will be based upon what
is known about the operations that were conducted in the room during and
prior to the current incident. In each situation, the aerosols will be
allowed to dissipate or settle before entry (approximately 30 minutes).
The following clothing requirements apply to these situations:
(a) BL-1. (1) Gloves.
(2) Outer complete covering such as a pair of coveralls.
(3) Shoe covers, provided shoes, or safety shoes or boots.
(4) Eye protection (maintenance only).
(b) BL-1 LS. The same as described in section 627.30(a) with the
following additions:
(1) An impervious apron.
(2) Impervious boots.
(c) BL-2 and toxins. (1) Gloves.
(2) Full outer covering such as a coverall.
(3) Shoe covers, provided shoes, or safety shoes or boots
(maintenance).
(4) An approved half-face or full-face respirator with HEPA filters
(worn).
(5) Eye protection.
(6) An impervious apron (not required for entry only).
(d) BL-2 LS. The same as Sec. 627.30(c) with the addition of
impervious boots.
(e) BL-3 and BL-3 LS. (1) A complete change of clothing.
(2) Gloves.
(3) An approved full-face HEPA or HEPA plus charcoal filtered
respirator.
(4) An impervious apron (not required for entry only).
(5) Impervious boots.
(6) Head cover.
(f) BL-4.
(1) A full change of inner clothing.
(2) An inner pair of gloves.
(3) A one-piece positive pressure suit as described in
Sec. 627.31(g), or a one-piece Xsuit with an approved positive pressure
self-contained breathing apparatus (SCBA) and a supplied-air respirator
(SAR) or both (see Sec. 627.31(f)).
(4) Appropriate gloves fitted to the suit.
(5) Impervious boots fitted over the suit.
Sec. 627.31 Specific requirements for individual PPE items.
(a) Aprons. Simple plastic or rubber aprons.
(b) Boots. When boots must be worn with an apron, the apron should
cover
[[Page 554]]
the boot tops sufficiently so that liquids splashed on the apron will
not run into the boots.
(c) Eye and face protection. Eye protection will meet or exceed the
requirements of OSHA found in the 29 CFR 1910.133 and will be worn at
all times when required. Special eye wear may be required around
ultraviolet (UV) light source.
(d) Gloves. (1) No one glove will be satisfactory for all
applications. Gloves are fabricated in a wide assortment of materials.
The type of glove selected will depend upon the specific activity. The
various activities in biocontainment facilities call for gloves to
protect against etiologic agents in situations where micro-manipulations
are required and excellent tactile feed-back through gloves is
important, gloves for handling hot glassware and cryogenic materials,
and gloves to protect against animal bites, toxic substances, chemical
carcinogens, solvents, acids, and caustics. Many of these requirements
call for gloves distinctly different from gloves suitable for the other
hazards. As a result, the SOP for each operation should address these
hazards and specify the appropriate glove required for each operation.
Consult MSDSs, manufacturer glove charts, and the safety officer to
determine the correct glove type needed.
(2) Before donning a pair of gloves, examine them closely to
ascertain that they are in serviceable condition. Check for rips and pin
holes. Gloves should over-wrap the cuff and lower sleeve of the
laboratory garment.
(3) Operations in open-front biological safety cabinets should be
planned so that once the operator has inserted gloved hands into the
cabinet, he or she does not have to withdraw them from the cabinet until
the work has been completed. If gloves become visibly contaminated, they
will be removed and decontaminated. Additional gloves should be
available so that work can continue. When wearing gloves for an extended
period, change them periodically or decontaminate them. Individual SOPs
will designate the appropriate period based upon the hazards.
(4) Gloves will be removed before going from one level of
containment to another (remove gloves in a safety cabinet before
removing your hands from the cabinet). Take care to ensure that skin is
not touched with the outer surface of contaminated or potentially
contaminated gloves when they are removed. Gloves will be placed in
suitable decontaminant when they are removed. Disposable gloves will be
placed in a covered container for decontamination or disposal.
(5) Gloves that are a part of a biological safety cabinet system
will be examined initially, after each sterilization of the biological
safety cabinet system, and at least annually for leaks using the soap
bubble test, followed by the halo-carbon test. Gloves will be tested
while still attached to the cabinet.
(6) Sterilization of nondisposable gloves either before use or
before reuse is usually done with ethylene oxide or formaldehyde gas.
Sterilized gloves must be aerated in flowing sterile (filtered) air at
21 deg.C or higher for a minimum of 24 hours prior to use to prevent
skin burns and irritation from residual decontaminants.
(e) Laboratory clothing. Users will check clothing before wearing
it, to ensure that it is free from defects that would compromise its
usefulness. Laboratory clothing (except BL-1) will be decontaminated
before being released for laundering by untrained or unprotected
personnel. Protective laboratory clothing that requires the wearer to
pull it over the head will not be used. Laboratory clothing will meet
OSHA requirements found in the 29 CFR 1910.132.
(f) One-piece suits. One-piece suits with a respirator under the
suit are not used to any great extent except in certain emergencies. The
respirators used with these are supplied air by an approved positive
pressure SCBA or SAR. Respirators will be of the pressure-demand or
constant flow type. The air provided will meet OSHA requirements found
in the 29 CFR 1910.134, the requirements of Grade D breathing air as
specified in the Compressed Gas Association pamphlet G-7.1 and American
National Standards Institute (ANSI) Z86.1-1973. When used in an area
that
[[Page 555]]
does not have a chemical shower to decontaminate the suit, a
decontamination station will be set up for this purpose. Suits
maintained for emergency use will be inspected at least quarterly and
respiratory equipment will be inspected monthly.
(g) One-piece positive pressure suits. A life-support system will be
provided with alarms and emergency backup breathing tanks. The air
provided will be HEPA-filtered meeting OSHA requirements found in the 29
CFR 1910.134, the requirements of Grade D breathing air as specified in
the Compressed Gas Association pamphlet G-7.1 and ANSI Z86.1-1973. A
HEPA-filter will be in-line between the disconnect on the suit and the
breathing space in the suit. When these are used in other than an
emergency situation, a chemical shower must be provided to decontaminate
the surfaces of the suit as the worker leaves the containment area.
Suits will be inspected before each use to check for indications of
significant wear or leakage. The suits will be worn with impervious
boots over the foot area of the suit and the outer gloves will be
attached over the hand portion.
(h) Respiratory protection equipment. (1) Respirators and their use
will be approved by the safety officer. The selection will be based on
the conditions of the activities and the risks involved. In general,
National Institute for Occupational Safety and Health (NIOSH) approved
respirators that use aerosol filters for dusts and fumes having a
Threshold Limit Value (TLV) of less than 0.05 mg/m3 have been
found acceptable for use in microbiological laboratories. Alternatively,
the Army M-17 or M-9 masks may be used. Air-supplied hoods are used in
situations where greater respiratory protection is required without the
need for body protection. One-piece suits are used when total body and
respiratory protection are required.
(2) When respirators are used, a respirator protection program will
be established that conforms to AR 11-34 and OSHA standards in the 29
CFR 1910.134. In general, a medical authority will designate who is to
wear respirators, they will be fitted by individuals trained in their
use and limitations, and wearers will be responsible for the proper
storage and regular inspection of their assigned respirators. Air-
purifying respirators will not be worn in oxygen deficient environments.
(3) Reusable respirators that have been worn in a contaminated area
will be decontaminated before reuse. At the end of each workday when a
respirator has been worn in an area where it was required, the wearer
will wipe it down with an appropriate liquid decontaminant. A damp cloth
soaked in the decontaminant, with the excess liquid squeezed out, will
be used for the wipe-down process, taking care to ensure that all
crevices are reached. The respirator will be rinsed with clean, warm
water. Visibly contaminated respirators will be decontaminated and
discarded.
(4) Respirator programs will comply with AR 385-10 and AR 11-34.
(i) Shoes. All shoes specially issued for use in controlled access
areas should be identified so that they can be segregated from other
areas. Safety shoes or boots meeting OSHA requirements stated in the 29
CFR 1910.134 will be issued wherever heavy items or corrosive chemicals
are handled. These will be sterilized appropriately after visible
contamination. In certain situations (excluding BL-4 operations), it is
desirable to wear disposable booties over street shoes, especially when
product protection is required.
Subpart E--Decontamination and Disposal
Sec. 627.32 Introduction.
All material or equipment that is potentially contaminated with
etiologic agents must be rendered nonhazardous before disposal. This
chapter describes the acceptable physical and chemical decontamination
methods and the general applicability of each. In general, all
infectious materials and all contaminated equipment or apparatus will be
sterilized before being washed and stored or discarded.
Sec. 627.33 Methods of decontamination.
(a) Autoclave. The use of wet heat is the most dependable procedure
for destroying all forms of microbial life. An autoclave employs
saturated steam under a pressure of approximately 15
[[Page 556]]
pounds per square inch (psi) to achieve a chamber temperature of at
least 121 deg.C for a minimum of 15 minutes. The time is measured after
the temperature of the material being sterilized reaches 121 deg.C.
Other combinations of temperature and pressure (some of which are
dependent on the equipment used) can be used to accomplish sterilization
provided that the efficacy of sterilization is validated as described
below. The most critical factor in ensuring the reliability of this
sterilization method, other than proper temperature, is preventing
entrapped air that is not replaced by stem. Material to be autoclaved
must come in contact with steam and heat and, as a result, it may be
necessary to add water to a load of waste to aid in the formation and
penetration of steam. Autoclaves use either a steam-activated exhaust
valve that reamins open during the replacement of air by live steam
until the steam triggers the valve to close, or a pre-cycle vacuum to
remove air prior to steam introduction.
(b) Sterilization will be verified using biological indicators (for
example, Bacillus stearothermophilus spores) at locations throughout the
autocalve, to include placement in the center of test loads, when the
autoclave is first put into service, and after any maintenance or
repairs. The primary means of verifying routine sterilization will be
through using chemical indicators (for example, autoclave tape or
labels) at locations throughout the autoclave. In addition each
autoclave will be equipped with a permanent means to record time and the
temperature of each operational event as a means of ensuring
sterilization. The type of materials being handled must be reviewed and
standard conditions for sterilization of each established. As a guide,
the manufacturer's manual for the autoclaves will be consulted as a
starting point in establishing these conditions. Treatment conditions to
achieve sterility will vary in relation to the volume of material
treated, the contamination level, the moisture content, and other
factors that should be considered and which may cause the times to
lengthen. In each case, the conditions will be established based on
tests which verify that the conditions selected are effective. In
addition to being effective from viable agents, autoclaving effectively
inactivates most protein toxins.
(c) Dry heat. Dry heat requires longer times or higher temperatures
or both than does wet heat. If used, the specific sterilization times
and temperatures must be determined for each type of material being
sterilized. In general, sterilization by dry heat can be accomplished at
169-170 deg.C for periods of 2 to 4 hours. Higher temperatures reduce
the time requirements. The heat transfer properties and spatial relation
or arrangement of materials in the load are critical in ensuring
effective sterilization.
(d) Liquid disinfectants. Liquid disinfectants may be used in
surface treatment, in dip tanks, and, at sufficient concentration, as
sterilants of liquid waste for final disposal. If liquid disinfectants
are used, they must have been shown to be effective against the
organisms present. Important considerations include: temperature, time
of contact, the negative logarithm of hydrogen ion concentration (pH),
concentration and state of dispersion, penetrability, and reactivity of
organic material at the site of application. Small variations in these
factors may make large differences in the effectiveness of disinfection,
so complete reliance should not be placed on liquid disinfectants when
the end result must be sterility. If evidence of efficacy under the
proposed procedures has not been reported previously, preliminary
studies to verify the efficacy of liquid disinfectants must be
conducted. Such studies may include attempts to recover and quantitate
the agent in question from liquid or swab samples, or sealed patches, by
animal inoculation, plaque assay, agar or broth cultivation, and similar
methods, following controlled decontamination under the same
experimental conditions envisioned for the proposed studies.
(1) Alcohol. Ethyl or isopropyl alcohol at the concentration of 70-
85 percent by weight will denature proteins but is slow in its
germicidal action. Alcohols are effective disinfectants for lipid-
containing viruses. These alcohols exhibit no activity against bacterial
spores.
[[Page 557]]
(2) Phenolic compounds. These are effective disinfectants against
vegetative bacteria, including Mycobacterium tuberculosis, fungi, and
lipid-containing viruses. The phenolics are not effective against
bacterial spores or non-lipid-containing viruses. The concentrations
used will be in accordance with the manufacturer's recommendations.
(3) Formaldehyde solutions. Formaldehyde in solution at a
concentration of 8 percent (formalin) is effective against vegetative
bacteria, spores, and viruses. It loses considerable disinfectant
activity below room temperature. Due to the toxic properties of
formaldehyde, the use of formalin is restricted to surfaces or materials
that are contained within appropriate engineering controls.
(4) Quaternary ammonium compounds. These cationic detergents are
strongly surface-active. They lose effectiveness in the presence of
proteins and are neutralized by anionic detergents, such as soap. At low
concentrations, they are bacteriostatic, tuberculostatic, sporostatic,
fungistatic, and algistatic. At medium concentration, they are
bactericidal, fungicidal, algicidal, and virucidal against lipophilic
viruses. They are not tuberculocidal, sporicidal, or virucidal against
hydrophilic viruses, even at high concentrations. The manufacturer's
recommended dilution will be used.
(5) Chlorine. Sodium hypochlorite is normally used as a base for
chlorine disinfectants. Free available chlorine is the active ingredient
and, at concentrations of at least 2,500 parts per million (ppm) (0.25
percent), is a disinfectant that is active against most microorganisms
and bacterial spores. Chlorine solutions at 2.5 percent free available
chlorine are effective against most toxins. Chlorine solutions lose
strength if exposed to air, so fresh solutions must be prepared whenever
the free chlorine content falls below desired minimums.
(6) Iodine. The characteristics of chlorine and iodine are similar.
Iodophor compounds with 1,600 ppm free available iodine provide a
relatively rapid inactivation of all microorganisms, including some
bacterial spores. A commonly available iodophor is Wescodyne. The
manufacturer of Wescodyne recommends a range of dilution form 1 to 3
ounces per 5 gallons of water, giving a solution containing from 25 to
75 ppm of free iodine. At these concentrations, available iodine may be
rapidly taken up by any extraneous protein present and will not be an
effective sporocide. A solution providing 1,600 ppm iodine is
recommended for hand washing or for use as a sporocide.
(7) Mercurials. Although the mercurials exhibit good activity
against viruses, they are toxic and are not recommended for general use.
They have poor activity against vegetative bacteria and are totally
ineffective sporicides. The dilution recommendations stated by the
manufacturer will be followed.
(e) Vapors and gases. Formaldehyde, ethylene oxide, peracetic acid,
beta-propiolactone, methyl bromide, and glutaraldehyde have all been
used successfully as space sterilants where they can be employed in
closed systems and with controlled conditions of temperature and
humidity. Of these, methyl bromide, beta-propiolactone, and
glutaraldehyde are not recommended because of their toxic properties.
Peracetic acid can readily decompose with explosive violence in a
concentrated state and must be used only in a diluted state and with
extreme care. Formaldehyde and ethylene oxide are both regulated by OSHA
for their potential human carcinogenicity, but do have permissible
exposure levels (unlike beta-propiolactone, for example) and can be used
safely under controlled conditions.
(1) Formaldehyde. Formaldehyde gas is, in general, the chemical of
choice for space disinfection. Biological safety cabinets and associated
effluent air-handling systems and air filters, incubators, laboratory
rooms, buildings, or other enclosed spaces can be disinfected with
formaldehyde. The procedures found in appendix E of the National
Sanitation Foundation Standard Number 49 will be followed for the
disinfection of biological safety cabinets. Other enclosures or areas
will be disinfected by following the same principles. To disinfect
rooms, the generation of formaldehyde gas from heating powdered or flake
paraformaldehyde is
[[Page 558]]
the preferred method. When area decontamination is performed, use 0.3
grams of paraformaldehyde for each cubic foot of space to be treated.
The room or area must be above 70 deg.F, the relative humidity above 70
percent, and the exposure time at least 2 hours (overnight is
preferred). After the required time for disinfection, the room must be
cleared of the formaldehyde gas (a small room with nonporous surfaces
and no materials or equipment in the room can be cleared of all
detectable formaldehyde by aeration for one hour, while larger areas
with equipment in them may take a full day). Before formaldehyde is used
as a space disinfectant, the area to be treated must be surveyed to
ensure that there are no open containers of any acidic solution
containing chloride ion in order to prevent the possible formation of
bis (chloromethyl)ether, a human carcinogen. Specific OSHA requirements
for posting of rooms and equipment, personnel protection, and other
requirements are found in 29 CFR 1910.1048.
(2) Ethylene oxide (EtO). EtO sterilization will only be conducted
in a sterilizer designed for that purpose and designed to maintain
potential exposure levels below the current OSHA standard. EtO is
effective against all microorganisms, including spores, molds,
pathogenic fungi, and highly resistant thermophilic bacteria. All
materials to be used in contact with human skin (for example, clothing,
shoes, masks, adhesive tape) must be aerated for at least 24 hours after
sterilization and prior to use. Concentrations of 500 to 1000 ppm are
required for sterilization. Specific OSHA requirements for the use of
ethylene oxide are found in 29 CFR 1910.1047.
(f) UV Radiation. UV radiation at a wave length of 253.7 nanometers
is a practical method for inactivating airborne virsuses, mycoplasma,
bacteria, and fungi. The usefulness of UV radiation on exposed surfaces
is limited by its low penetrating power. UV radiation shall only be
relied upon to sterilize surfaces when conventional methods, such as
autoclaving or the use of liquid disinfectants, would make the product
unusable. An example is data sheets that must be brought out of a
biocontainment facility. The UV intensity must be at least 40
microwatts/cm 3 on the surface to be treated. Single sheets
of paper may be treated by exposing them to this radiation for a minimum
of 15 minutes. A calibrated photoelectric UV intensity meter, capable of
measuring UV radiation at a wave length of 253.7 nanometers, will be
used whenever a new UV source is installed, and quarterly thereafter, to
ensure the UV source is providing at least 40 microwatts/cm 3
at the work surface. Bulbs should be cleaned routinely to remove any
accumulated dust and prolong bulb performance and assure proper energy
output. Protective eye wear and clothing may be necessary when working
around UV radiation.
Sec. 627.34 Disposal.
Inactivation is the first step in the disposal of etiologic agents
or materials that are potentially contaminated with them. All
contaminated or potentially contaminated materials must be effectively
disinfected or sterilized by an approved procedure discussed in
Sec. 627.33. After decontamination, reusable items, such as clothing or
glassware, may be washed with other uncontaminated or decontaminated
items.
(a) Combustible items. Combustible disposable items should be bagged
and incinerated in an appropriate approved incinerator or otherwise
disposed of in accordance with State and local regulations.
(b) Noncombustible disposable items. Items will be packaged as
stated in Sec. 626.34(e) and disposed of by a licensed waste hauler.
(c) Equipment. Equipment that cannot be autoclaved will be
decontaminated by gaseous sterilization or with a suitable liquid
disinfectant. Such equipment will be certified as decontaminated by the
safety officer.
(d) Waste. Materials generated, such as solvents, acids, chemical
carcinogens, radioactive isotopes, medical waste, or dead animals must
be decontaminated, packaged, and then disposed of in accordance with
EPA, NRC, local, State, and Federal regulations.
[[Page 559]]
(e) Mixed waste. When two or more hazardous materials are mixed
together, the mixture will be decontaminated and disposed of in
accordance with EPA, NRC, State, and Federal regulations for the
mixture, or for the most hazardous material.
(f) Packaging. Solid waste will be placed in cans, sturdy bags, or
boxes. Rigid, puncture-resistant, sealable containers will be used for
packaging ``sharps.'' When wet materials are packaged for disposal, the
materials will be placed in a leak-proof container. Heavy waste will be
placed in rigid containers ensuring that the burst strength of the
container is not exceeded.
(g) Labeling. A method of verifying that all items prepared for
disposal have been decontaminated will be established for etiologic
agent wastes. Mixed waste will be labeled as appropriate to indicate the
hazards that must be addressed after decontamination.
(h) Recordkeeping. A manifest will be initiated and maintained,
where required, to record the disposition and transfer of waste.
Applicable Federal, State, and local ordnances will be followed.
Subpart F--Importation, Shipment, and Transport of Etiologic Agents
Sec. 627.35 Introduction.
The CDC of the Public Health Service (PHS), the United States
Department of Agriculture (USDA), the Food and Drug Administration
(FDA), the Department of Transportation (DOT), the United States Postal
Service and the International Air Transport Association (IATA) regulate
the importation, shipment, and transportation of etiologic agents. This
chapter outlines the minimum administrative requirements the commander
or institute director are to follow and gives sources for information on
the requirements for importation, packaging, labeling, and shipment of
etiologic agents.
Sec. 627.36 Administration.
The commander or institute director will establish the following
controls to ensure that etiologic agents are transported with proper
authorization, controls, and procedures:
(a) Institute policies will be established in writing to ensure that
before etiologic agents are acquired or shipped--
(1) The division chief responsible for the area where work with
etiologic agents is to be conducted approves all acquisitions or
shipments.
(2) The safety officer is informed in writing of the type and amount
of any BL-4 or USDA-restricted etiologic agent (listed in HHS
publication No. (NIH) 88-8395 or current edition) being received, and
the estimated date of arrival.
(3) The recipient of all etiologic agents shipped from an institute
will be documented.
(4) The commander or institute director approves all acquisitions
and shipments of BL-4 or USDA-restricted etiologic agents.
(5) The commander or institute director approves all requests for
shipments to or from foreign countries and to individuals not affiliated
with an institution or agency (for example, physicians in private
practice).
(6) The Office of The Surgeon General, United States Army, or the
Commander, United States Army Materiel Command (AMC) approves the
initial acquisition and use of all reference stocks of etiologic agents
and transfers between Army RDTE activities in accordance with AR 70-65.
(7) There is full compliance with the regulatory requirements
referenced in Secs. 627.37, 627.38, 627.39 and 627.40.
(8) The following information regarding the recipient and the
intended use of BL-4 and USDA-restricted animal pathogens, will be kept
on file for 10 years. This information will also be kept for all
shipments to or from foreign countries and to individuals not affiliated
with an institution or agency (for example, physicians in private
practice).
(i) The requester's name and address.
(ii) The type and amount of the etiologic agent to be sent.
(iii) The qualifications of the recipient of the etiologic agent.
(iv) The intended use of the etiologic agent.
[[Page 560]]
(v) A statement indicating that the agent is not for human use.
(b) Etiologic agents assigned to biosafety level 1, 2, or 3,
approved for shipment, and properly labeled and packaged may be shipped
by commercial cargo carriers.
(c) All etiologic agents assigned to BL-4 or USDA-restricted animal
pathogens approved for shipment and properly packaged, will be
accompanied by a designated courier, or under close supervision of a
responsible party who will monitor aspects of the shipment, ensuring
that required transfers have been completed and documented and final
receipt has been accomplished and acknowledged.
Sec. 627.37 Importation directives.
Importation of etiologic agents is subject to the Public Health
Service Foreign Quarantine Regulations (42 CFR 71.156). Examples of
permits authorizing the importation or receipt of regulated materials
and specifying conditions under which the etiologic agent is shipped,
handled, and used are contained in appendix E to this part.
Sec. 627.38 Shipment directives.
Shipping unmarked and unidentified etiologic agents is prohibited.
Etiologic agents will be packaged, labeled, and shipped according to the
requirements found in the Interstate Shipment of Etiologic Agents
Regulations (42 CFR part 72) and its amendments. The USDA regulations in
9 CFR parts 102 through 104, 122 and the FDA regulations in 21 CFR parts
312 and 600 through 680 will also be followed as applicable. Packaging
and labeling requirements for interstate shipment of etiologic agents
are summarized and illustrated in appendix D. Permits authorizing the
shipment of regulated materials and specifying conditions under which
the etiologic agent is shipped, handled, and used are contained in
appendix E to this part.
Sec. 627.39 Transportation directives.
The packaging and labeling requirements cited above must be followed
for the local transport of etiologic agents and diagnostic specimens by
courier or by other delivery services. Similar requirements and
restrictions applicable to the transport of etiologic agents, diagnostic
specimens, and biological products by all modes of transportation (that
is, air, motor, rail, and water) are imposed by the Department of
Transportation (49 CFR part 173), IATA ``Dangerous Goods Regulations,''
the Air Transport Association ``Restricted Articles Tariff 6-D,'' the
International Civil Aviation Organization (ICAO), Postal Bulletin No.
21246 ``International Mail-Hazardous Materials,'' 39 CFR, and, the
Domestic Mail Manual. When shipments exceed 4 liters, the requirements
found in AR 740-32 will be followed.
Sec. 627.40 Additional requirements.
Additional requirements for importation, shipment, and
transportation of infectious agents and hazardous materials that must be
followed are contained in the following directives:
(a) AR 40-12, Medical and Agricultural Foreign and Domestic
Quarantine Regulations for Vessels, Aircraft, and Other Transports of
the Armed Forces.
(b) AR 70-65, Management of Controlled Substances, Ethyl Alcohol,
and Hazardous Biological Substances in Army Research, Development, Test,
and Evaluation Facilities.
Sec. 627.41 Sources for further information on shipment of etiologic agents.
(a) Guide for Transportation of Hazardous Materials, Vol. 4(1),
February 10, 1975. Copies are obtainable from the Office of Research
Grants Inquiries, NIH, Department of Health and Human Services, 5333
Westbard Avenue, Bethesda, MD 20205.
(b) The CDC, Office of Biosafety, 1600 Clifton Road N.E., Atlanta,
Georgia 30333. Telephone (404) 639-3883, or FTS: 236-3883.
(c) The American Type Culture Collection (ATCC), Packaging and
Shipping of Biological Materials at ATCC. Copies may be obtained from
the ATCC, 12301 Parklawn Drive, Rockville, MD 20852. Phone (301) 881-
2600.
(d) National Committee for Clinical Laboratory Standards (NCCLS),
Procedures for the Domestic Handling and Transport of Diagnostic
Specimens and
[[Page 561]]
Etiologic Agents, (H5-A2), Second edition. Vol. 5, No. 1. Copies are
obtainable from the NCCLS, 771 East Lancaster Avenue, Villanova, PA
19085.
Subpart G--Facilities
Sec. 627.42 Introduction.
The design of the facility is important in providing a secondary
barrier to protect individuals inside and outside the facility. Because
the hazards presented by various organisms and materials vary, the
requirements for the facility will vary accordingly. The minimum
facility requirements for the various biosafety levels and toxins are
described below. The biosafety levels correspond to those described in
the HHS Publication Biosafety in Microbiological and Biomedical
Laboratories (HHS No. (NIH) 88-8395), while the large-scale biosafety
levels were adapted from those described in the NIH Guidelines for
Research Involving Recombinant DNA Molecules.
Sec. 627.43 Biosafety level 1.
(a) Laboratories. Each laboratory used for this level will, as a
minimum, have the following features:
(1) A sink for handwashing.
(2) Work surfaces that are impervious to water and resistant to
acids, alkalis, organic solvents, and moderate heat.
(3) Fly screens on any windows that can be opened.
(4) Furnishings and surfaces that are sturdy and designed to be
easily cleaned.
(5) Spaces between furnishings and equipment that are accessible for
cleaning.
(b) Animal facilities. Each room will have the following features:
(1) Design and construction to facilitate cleaning and housekeeping.
(2) A sink for handwashing within the facility.
(3) Fly screens on any windows that can be opened.
(4) Ventilation designed so that the direction of airflow in the
animal facility is inward, with the exhausted air discharged to the
outside without being recirculated.
(5) Self-closing doors that open inward.
Sec. 627.44 Biosafety level 2.
(a) Laboratories. Each laboratory used for this level of hazard will
have, in addition to the requirements stated in Sec. 627.43(a), the
following:
(1) An autoclave available.
(2) Containment equipment necessary for the operations unless the
safety officer approves the use of a compensatory level of personal
protective equipment.
(3) An eyewash available near the laboratory.
(b) Animal facilities. In addition to the requirements stated in
Sec. 627.43(b), facilities will include--
(1) A sink for handwashing in each room where animals are housed.
(2) An autoclave available in the building.
(3) Appropriate containment equipment unless the safety officer
approves the use of a compensatory level of personal protective
equipment.
Sec. 627.45 Biosafety level 3.
(a) General requirements. Each suite used as a laboratory or in
which infected animals are housed will, as a minimum, have the following
features:
(1) Physical separation from areas which are open to unrestricted
traffic.
(2) All entrances to each laboratory or animal room from the
nonlaboratory access corridors will be through two sets of doors. A
change room or airlock may be incorporated between the doors.
(3) The interior surfaces of walls, floors, and ceilings will be
water resistant so that they may be easily cleaned.
(4) All penetrations into the walls, floors, and ceilings should be
sealed or capable of being sealed to facilitate decontamination.
(5) A foot, elbow, or automatically operated sink will be located
near the exit door to each laboratory or animal room.
(6) An autoclave should be in each laboratory or animal room and
will be available to the facility.
(7) A ventilation system that will--
(i) Create directional airflow that draws air into the laboratory
through the entry areas.
(ii) Not recirculate laboratory air.
[[Page 562]]
(iii) Discharge the exhaust air from the laboratory to the outside
and disperse the exhaust air away from occupied areas and air intakes.
(iv) Exhaust the HEPA-filtered air from Class I or II biological
safety cabinets or other primary containment devices directly to the
exterior of the laboratory or through the building exhaust system.
Exhaust air from the cabinets may be recirculated within the laboratory
if the cabinet is tested and certified at least every 12 months. If the
filtered cabinet exhaust is discharged through the building exhaust
system, it will be connected to this system in a manner (for example,
thimble unit connection) that avoids any interference with the air
balance of the cabinets or the building exhaust system.
(8) All windows to the facility will be sealed shut.
(9) Appropriate biological safety cabinets or other specialized
containment equipment will be provided.
(10) Any vacuum line in the facility will have a HEPA filter and
liquid disinfectant trap.
(11) Bench tops that are impervious to water and resistant to acids,
alkalis, organic solvents, and moderate heat.
(12) Furnishings that are sturdy and spaces between benches,
cabinets, and equipment that are accessible for cleaning.
(13) An eyewash available in or near the laboratory.
(b) Additional animal facility requirements. In addition to the
requirements given in Sec. 627.44(b) and 627.45(a), all doors to the
animal rooms will open inward and be self-closing.
Sec. 627.46 Biosafety level 4.
The engineering controls within the facility must provide absolute
biological containment. All procedures with etiologic agents requiring
this biosafety level of facilities, equipment, and procedures must be
conducted either in Class III biological safety cabinets, or in a
facility that is designed for the use of a personal positive pressure
suit as described in Sec. 627.46(b) in conjunction with Class I or II
biological safety cabinets.
(a) General requirements. The facility will have the following
features:
(1) A separate building or a clearly demarcated and isolated area
within a building which incorporates positive personnel control for
access.
(2) All entrances from access corridors incorporate an inner and
outer change room.
(3) Inner and outer change rooms separated by a shower facility.
(4) A double-doored autoclave, fumigation chamber, or ventilated
airlock for passage of all items which do not enter the facility through
the change room.
(5) Interior surfaces of walls, floors, and ceilings resistant to
water and chemicals to facilitate cleaning and disinfecting.
(6) Walls, floors, and ceilings of the facility constructed to form
a sealed internal shell which facilitates fumigation and is animal and
insect proof.
(7) All penetrations into the walls, floors, and ceilings sealed.
(8) All liquid drains in the facility connected directly to a liquid
waste decontamination system.
(i) Holding tanks collecting waste from sinks, biological safety
cabinets, floors, and autoclave chambers provide decontamination by heat
treatment.
(ii) Holding tanks collecting waste from shower rooms and toilets
provide decontamination by heat or chemical disinfectant methods.
(9) Sewer and other ventilation vents contain in-line HEPA filters.
(10) Internal facility appurtenances (for example, light fixtures,
air ducts, and utility pipes) arranged to minimize the horizontal
surface area on which dust can settle.
(11) A foot, elbow, or automatically operated handwashing sink
located near the exit door to each laboratory or animal room.
(12) Self-closing and lockable access doors.
(13) A ventilation system that--
(i) Is dedicated to the facility and provides fresh air meeting
American Society of Heating, Refrigerating, and Air Condition Engineers,
Inc. (ASHRAE) Standard 62.
(ii) Maintains a negative pressure differential and assures flow
inward from areas outside of the facility toward areas of highest
potential risk.
[[Page 563]]
(iii) Has manometers or magnehelic gauges to provide, sense, and
display pressure differentials between adjacent areas maintained at
different pressure levels. An alarm will sound when the pressures fall
below acceptable levels.
(iv) Has the air supply and exhaust interlocked to ensure that
exhaust failure or reduction will not allow the air pressure in the area
to become positive to the adjacent areas.
(v) Does not recirculate exhaust air.
(vi) Is HEPA-filtered and discharged to the outside, dispersing the
exhaust air away from occupied areas and air intakes.
(vii) Has the HEPA filters on the exhaust located as near to the
rooms as is practicable.
(viii) Has the filter chambers designed to allow in-place
decontamination before the filters are removed and to facilitate
certification testing.
(ix) Contains prefilters and HEPA filters in the air supply system
to protect the supply air system should air pressures become unbalanced.
(x) Exhausts the HEPA-filtered air from Class I or II biological
safety cabinets directly into the laboratory or to the exterior of the
building. If the HEPA-filtered exhaust from these cabinets is
recirculated, the cabinets are tested and certified every 6 months. If
the filtered cabinet exhaust is discharged through the building exhaust
system, it will be connected to this system in a manner (for example,
thimble unit connection) that avoids any interference with the air
balance of the cabinets or the building exhaust system.
(xi) Passes the treated exhaust air from Class III biological safety
cabinets through two sets of HEPA filters in series to the exterior of
the facility through the laboratory exhaust air system.
(14) Windows (if present) sealed shut and breakage resistant.
(15) Has a double-doored autoclave for decontaminating materials
passing out of the facility. The autoclave door that opens to the area
external to the facility is sealed to the outer wall and automatically
controlled so that it can only be opened after the autoclave
sterilization cycle has been completed.
(16) Has a pass-through dunk tank, fumigation chamber, or an
equivalent decontamination method for materials and equipment that
cannot be autoclaved.
(17) Has central vacuum systems (if present) that--
(i) Do not serve areas outside the facility.
(ii) Have an in-line HEPA filter placed as near as practicable to
each use point or service cock.
(iii) Have filters designed to allow in-place decontamination and
replacement.
(18) Liquid and gas services to the facility provided with
protective devices that prevent backflow.
(b) Additional requirements for personal positive pressure suit
areas. If personal positive pressure suits are worn in lieu of using
Class III biological safety cabinets for containment, a special suit
area will be provided. The suit area will provide the following, in
addition to the requirements stated in Sec. 627.46(a):
(1) An exhaust system dedicated to that area that provides
filtration by two sets of HEPA filters installed in series. This system
will be backed up by a duplicate filtration unit, exhaust fan, and an
automatically starting emergency power source. The ventilation system
will maintain the suit area under negative pressure relative to the
surrounding areas.
(2) An entry area consisting of an airlock fitted with airtight
doors.
(3) A chemical shower to decontaminate the surface of the personal
positive pressure suit upon exit.
(4) An air supply and distribution system to support the life
support system of the personal positive pressure suits.
(5) Emergency lighting and communications systems.
(6) Sealed penetrations into the internal shell of the area.
(7) A double-doored autoclave to decontaminate waste materials to be
removed from the suit area.
(c) Additional laboratory requirements. In addition to those given
in Sec. 627.45, if water fountains are provided, they will be foot
operated and located in the facility corridors outside the laboratory.
[[Page 564]]
(d) Additional animal facility requirements. In addition to those
requirements given in Sec. 627.45, all animal facility external doors
will be self-locking.
Sec. 627.47 Large-scale facilities.
The following requirements apply to facilities in which an
individual culture of viable etiologic agents exceed 10 liters:
(a) BL-1 LS. In addition to the laboratory requirements stated
Sec. 627.43(a), the exhaust gases removed from a closed system or other
primary containment equipment shall be treated by filters which have
efficiencies equivalent to HEPA filters or by other equivalent
procedures (for example, incineration) to minimize the release of viable
organisms.
(b) BL-2 LS. In addition to the requirements stated in
Secs. 627.44(a) and 627.47(a), these facilities will have--
(1) Rotating seals and other mechanical devices directly associated
with a closed system used to contain viable organisms shall be designed
to prevent leakage or shall be fully enclosed in ventilated housings
that are exhausted through filters which have efficiencies equivalent to
HEPA filters or through equivalent treatment devices.
(2) A closed system used to propagate and grow viable organisms
shall include monitoring or sensing devices that monitor the integrity
of containment during operations.
(3) Closed systems used for the propagation and growth of viable
organisms shall be tested operationally for integrity of the containment
features. The containment will be rechecked following modification or
replacement of essential containment features. Procedures and methods
used in the testing shall be appropriate for the equipment design and
for recovery and demonstration of the test organism. Records of tests
and results shall be maintained on file.
(c) BL-3 LS. The requirements stated in Secs. 627.45 and 627.57(b)
apply, and all closed systems and other primary containment equipment
used in handling cultures of viable organisms shall be located within a
controlled area which meets the requirements of a BL-3 facility plus the
following requirements:
(1) All utilities and service or process piping or wiring entering
the controlled area shall be protected against contamination.
(2) A shower facility shall be provided. This facility shall be
located near the controlled area.
(3) The controlled area shall be designed to preclude release of
culture fluids outside in the event of an accidental spill or release
from the closed systems or other primary containment equipment.
(4) The controlled area shall have a ventilation system capable of
controlling air movement. The movement of air shall be from areas of
lower contamination potential to areas of higher contamination
potential. If the ventilation system provides positive pressure supply
air, the system shall operate so as to prevent the reversal of air
movement or shall be equipped with an alarm that would be actuated if
reversal in the direction of air movement were to occur. The exhaust air
from the controlled area shall not be recirculated to other areas of the
facility. The exhaust air from the controlled area may be discharged to
the outdoors after filtration or other means of effectively reducing an
accidental aerosol burden, and dispersed clear of occupied buildings and
air intakes.
Sec. 627.48 Toxins.
General requirements for all facilities in which toxins are used are
as follows. Such facilities will--
(a) Have a ventilation system that provides three to six air changes
per hour, and that provides a directional airflow inward relative to the
access halls.
(b) Have a sink for handwashing.
(c) Have an eyewash available.
(d) Have bench tops that are impervious to water and resistant to
acids, alkalis, organic solvents, and moderate heat.
(e) Have furniture, furnishings, and surfaces that are sturdy and
designed to be easily cleaned.
(f) Be arranged so that items are accessible for cleaning.
(g) Have a quick-drench shower available within the facility.
[[Page 565]]
(h) A fume hood, biological safety cabinet, glove box, or equivalent
engineering control equipped with HEPA filters and with charcoal filters
if volatile materials are being used.
Subpart H--Engineering Controls
Sec. 627.49 Introduction.
As required by the OSHA and recommended by the American Industrial
Hygiene Association (AIHA) and the CDC, engineering controls and proper
microbiological techniques are the primary means of protecting personnel
who work with potentially hazardous biological materials. In situations
of potentially higher hazard, these engineering controls are
supplemented by personal protective clothing and equipment. Thus, the
engineering controls discussed in this chapter will be the primary means
of personnel and environmental protection when working with etiologic
agents. Because of the importance of these engineering controls, this
chapter contains not only requirements for the engineering and
construction of these controls, but also requirements for their
certification and continuous satisfactory performance. These will be
described for each engineering control.
Sec. 627.50 Class I biological safety cabinet.
(a) Description. The Class I biological safety cabinet (figure H-I
in appendix F to this part) is a ventilated cabinet for personnel
protection only. The cabinet provides an uncirculated inward flow of air
away from the operator. The exhaust is passed through a HEPA filter. It
may be discharged into the laboratory or vented out of the laboratory
and dispersed away from occupied spaces or air intakes. When the exhaust
is recirculated in a BL-2 or BL-3 facility, the cabinet must be tested
and certified annually. In a BL-4 facility, if the exhaust is
recirculated, the cabinet must be tested and certified semiannually.
(b) Uses. These cabinets are used if personnel protection against
the microorganisms is required; for modest quantities of volatile,
toxic, or radioactive chemicals (in concentrations and quantities
associated with biological systems) if vented to the outside; and when
sterility is not required. They are commonly used for housing tabletop
centrifuges, in the necropsy of small animals, and for changing animal
bedding.
(c) Prohibitions. This class of cabinet is not to be used when
sterility must be maintained. In addition, volatile, toxic, or
radioactive materials can not be used in this class of cabinet when the
exhaust air is not exhausted to the exterior.
(d) Certifications and requirements. (1) The inward air velocity on
these cabinets will be an average of 100 plus or minus 20 linear feet
per minute (lfpm). Each cabinet must be certified before use and
semiannually thereafter by a face velocity test. Additionally, smoke
tests will be performed annually to verify containment.
(2) The exhaust system will have a HEPA filter, which will be tested
initially upon installation, after repair or replacement, and every 2
years thereafter (except when required more often). Filters will be
certified to be 99.97 percent effective in capturing particulate matter
by a leakage test using mineral oil or other appropriate aerosol
dispersed as 0.3 micron droplets.
Sec. 627.51 Class II biological safety cabinet.
All Class II biological safety cabinets (figure H-II in appendix F
to this part) are ventilated cabinets for personnel and product
protection, having an open front with inward air flow for personnel
protection.
(a) Operating standards. (1) All of these cabinets must conform and
be certified to meet National Sanitation Foundation (NSF) Standard No.
49 revised, June 1987, for the applicable type of cabinet.
(2) After installation and before use, and annually thereafter, the
cabinets will be tested in accordance with NSF Standard No. 49 (latest
revision June 1987) as follows:
(i) Primary (required) tests--
(A) Velocity profile test.
(B) Work access opening airflow (face velocity) test.
(C) HEPA filter leak test.
[[Page 566]]
(D) Cabinet integrity test (soap bubble test) for cabinets with
positive pressure internal plenums.
(ii) Secondary (optional) tests--
(A) Vibration test.
(B) Electrical leakage and ground circuit resistance tests.
(C) Noise level test.
(D) Lighting intensity test.
(E) UV light intensity test.
(3) After repairs or alterations to the cabinetry or ventilation
system that affect the cabinet, the tests listed in Sec. 627.51(a)(2)
will be performed for the relevant parameters.
(4) The work access opening airflow (face velocity) test, as
specified in NSF Standard No. 49 (latest revision, June 1987), will be
performed to check that the cabinet is within specifications on an
annual basis for BL-1 and BL-2 and toxin use. This test will be
performed semiannually on cabinets used for BL-3 and BL-4 as well as for
work with dry forms of toxins.
(5) When the exhaust is recirculated in a BL-4 facility, the cabinet
must be tested and certified semiannually.
(b) Class IIA biological safety cabinets.--(1) Description. A Class
IIA biological safety cabinet is one in which typically 70 percent of
the air is recirculated within the cabinet and the exhaust passes
through a HEPA filter before discharge. The exhaust may be exhausted
into the room and positive-pressure contaminated ducts and plenums
within the cabinet are allowed. Type A cabinets shall have a minimum
calculated face velocity of 75 feet per minute (fmp).
(2) Uses. These cabinets are for working with low-to-moderate risk
biological samples and for protecting personnel against biological
material while providing a sterile atmosphere in which to handle the
material.
(3) Prohibitions. Materials that are toxic or volatile must not be
used in these cabinets.
(c) Class IIB1 biological safety cabinets.--(1)
Description. A Class IIB1 biological safety cabinet is one
that maintains a minimum average inflow of air of 100 plus or minus 20
lfpm and in which typically 30 percent of the air is recirculated. All
recirculated and exhausted air passes through two HEPA filters in
series. All contaminated internal ducts and plenums are under negative
pressure. Type B cabinets shall have a minimum calculated face velocity
of 100 fpm.
(2) Uses. When ultra-sterility is needed, these are the cabinets of
choice. The double filtration achieves a cleaner atmosphere. Minute
quantities of volatile, toxic, or volatile radioactive materials
coincidental to use in biological systems may also be used in these
cabinets.
(3) Prohibitions. More than minute quantities of toxic, volatile, or
radioactive materials must not be used in these cabinets.
(4) Additional certifications or requirements. None.
(d) Class IIB2 biological safety cabinets.--(1)
Description. A Class IIB2 biological safety cabinet is one
that maintains a minimum average of 100 plus or minus 20 lfpm inward
flow and in which all air is exhausted directly from the cabinet through
a HEPA filter without recirculation within the cabinet. All contaminated
ducts and plenums are under negative pressure. Type B cabinets shall
have a minimum calculated face velocity of 100 fpm.
(2) Uses. These cabinets are recommended when small quantities of
volatile, flammable, or toxic chemicals must be used coincidentally with
items requiring sterility.
(3) Prohibitions. While these cabinets do offer the greatest degree
of safety for volatile, toxic, and flammable chemical handling in a
sterile environment, they are not to be used in place of a fume hood to
prepare stock solutions of hazardous chemicals.
(e) Class IIB3 biological safety cabinets.--(1)
Description. A Class IIB3 biological safety cabinet is one
that meets all of the requirements of a Class IIB2 biological
safety cabinet except that it recirculates most (typically 70 percent)
of the air inside the cabinet. Type B cabinets shall have a minimum
calculated face velocity of 100 fpm.
(2) Uses. Minute amounts of nonflammable chemicals can be used
coincidentally with low-to-moderate risk biological agents.
(3) Prohibitions. Flammable materials and more than minute amounts
of toxic, radioactive, or volatile chemicals must not be used in these
cabinets.
[[Page 567]]
(4) Additional certifications or requirements. None.
Sec. 627.52 Class III biological safety cabinet.
(a) Description. These cabinets (figure H-III in appendix F to this
part) are totally enclosed, ventilated cabinets of gas-tight
construction. Operations are conducted through attached rubber gloves.
The supply of air is drawn into the cabinet through HEPA filters. The
exhaust air is treated by double HEPA filtration, or by HEPA filtration
followed by incineration, and is not allowed to recirculate within the
room.
(b) Uses. These cabinets provide the ultimate protection for
personnel. They are suitable for low, moderate, and high-risk etiologic
agents.
(c) Prohibitions. More than minute amounts of flammables must not be
used in these cabinets.
(d) Certifications and requirements. (1) These cabinets will have a
manometer or magnehelic gauge that indicates the negative pressure that
is maintained inside the cabinet. The pressure inside the cabinet should
be a minimum of 0.5 inches water gauge negative to the surrounding room.
(2) These cabinets will be pressure tested by the soap bubble or
halogen leak test as prescribed in NSF Standard No. 49, appendix B1
(latest revision, June 1987), and certified, when the HEPA filter units
are serviced.
Sec. 627.53 Fume hood.
Fume hoods in which etiologic agents are handled must use proven
technologies to provide optimal containment. Fume hood placement,
design, and capture testing requirements for use in designing new
laboratories can be found in the latest edition of Industrial
Ventilation, A Manual of Recommended Practices, published by the
American Conference of Governmental Industrial Hygienists.
(a) Description. Fume hoods are common chemical laboratory
furnishings designed to capture fumes from chemicals that are used
within them. Air is drawn through the opening and vented to the exterior
without recirculation.
(b) Uses. Fume hoods provide excellent containment for handling
hazardous chemicals.
(c) Prohibitions. Moderate risk biologicals and open containers of
dry forms of toxins must not be used in a fume hood without HEPA
filtration. Fume hoods should never be used when sterility is required.
(d) Certification and requirements. (1) Inward air flow will be an
average of 100 plus or minus 20 lfpm as measured at the face of the fume
hood. Proper function of laboratory hoods is not only a function of face
velocity. An evaluation of the total operating environment is necessary.
(2) When filters are required, they will be certified by the mineral
oil droplet (HEPA) or Freon (Charcoal) leak test as appropriate. Leakage
through the filters will be less than 0.05 percent for Freon and 0.03
percent for oil droplets when initially installed.
(3) Fume hoods will be provided with indicator devices to give a
warning should the ventilation system fail or if the hood face velocity
falls below an average of 80 lfpm
(4) Hood air flow will be certified when installed, when maintenance
is performed on the ventilation system, and semiannually thereafter.
Sec. 627.54 Glove box.
(a) Description. A glove box is an enclosure that provides a
positive barrier from liquids, solids, and chemical vapors. A glove box
has viewing ports and glove ports for access. The box maintains
personnel protection through solid barriers and maintenance of a
negative pressure relative to its surroundings.
(b) Uses. Glove boxes are used when extreme containment is needed
for highly toxic chemicals, especially for dry chemicals that can be
swept out of containers by the airflow in hoods.
(c) Prohibitions. Unventilated boxes must not be used with volatile
flammable materials and should be used with volatile toxic materials
unless dilution ventilation is provided.
(d) Additional certifications and requirements. (1) The glove box
will be maintained at a pressure of at least 0.25 inches water guage
less than its surroundings.
(2) The pressure differential will be indicated by a manometer or
magnehelic guage. Indicator devices
[[Page 568]]
will display a loss of pressure below 0.25 inches water guage.
(3) Gloves will be changed at appropriate intervals (dependent on
the box contents) to ensure they provide the protection needed.
(4) Inlets that provide dilution air will be protected by HEPA
filters.
Sec. 627.55 Ventilated balance enclosures.
(a) Description. A ventilated balance enclosure is a box that
surrounds a balance and has a small open area for access and handling
material in the front. Air is exhausted out the rear of the enclosure.
(b) Uses. A ventilated balance enclosure is used when containment of
a balance is required to weigh hazardous materials that have a low vapor
pressure (such as toxins). These enclosures are also used when it is
best to use the balance in other than a fume hood (due to the turbulence
and vibration) and when biological safety cabinets or glove boxes are
inappropriate or unavailable. Dry forms of toxins may be weighed in
these enclosures.
(c) Prohibitions. Very volatile or highly toxic volatile materials
must not be handled in ventilated balance enclosures unless they are
placed in closed containers in a properly functioning fume hood before
being transferred to the balance enclosure.
(d) Additional certifications or requirements. (1) The flow through
the openings in the enclosure will be at least 60 lfpm and must average
between 60 and 80 lfpm.
(2) Containment will be certified prior to first use and annually
thereafter by smoke tubes.
(3) The air flow will be certified initially and semiannually by
averaging readings taken from the face of the opening.
Sec. 627.56 Ventilated cage enclosures.
There are a number of cage-ventilated enclosures in which infected
animals may be housed at levels corresponding to the various classes of
biological safety cabinets. A brief description of four different types
of animal ventilated cages is given below. This is not a complete
description of all the different animal ventilated cages available. The
proper functioning of these will be tested initially, upon each
connection to exhaust sources, and at least annually. The inward flow
rates on the partial containment systems and pressure checks on the
total containment cages will be performed. Prior to selecting such
equipment, an evaluation of the function and the equipment should be
made, and the methods for testing and decontamination should be analyzed
and documented.
(a) Filter-top cages. Small laboratory animal polystyrene or
polycarbonate cage bottoms are fitted with a dome shaped glass fiber or
polyester filter cage cover. The dome shaped filters help reduce the
dissemination of aerosols, and the spread of infectious agents. Adequate
ventilation around cages fitted with a dome shaped filter is essential
since they may contain elevated ammonia and carbon dioxide levels, and
high temperature and humidity. Ventilation recommendations in the NIH
publication 86-23, 1985 ``Guide for the Care and Use of Laboratory
Animals'' will be followed.
(b) Forced ventilation cages. This is a small HEPA-filtered cage
connected to a centralized exhaust system. A minimum airflow of 0.03
m\3\ /min per cage is required. Ventilation rates may vary with the size
of the cage, and the number and type of animals being housed.
(c) Cubicle-type isolation cage. This is a partial containment unit
which holds several animal cages. This unit is a negative pressure HEPA-
filtered stainless steel cage. A minimum airflow of 0.3 m\3\ /min per
cage is required for a 0.24 m\3\ unit. Ventilation rates may vary with
the size of the cage and the number and type of animals being housed.
(d) Total containment cage. This unit is a negative pressure or
positive pressure HEPA-filtered stainless steel cage which has the
filters incorporated into the design. It is halogen gas-leak tight and
can be considered a Class III biological safety cabinet. A minimum
airflow of 0.3 m\3\ /min per cage is required for a 0.24 m\3\ unit.
Ventilation rates may vary with the size of the cage, and the number and
type of animals being housed.
[[Page 569]]
Sec. 627.57 Ventilated cage areas.
Ventilated cage areas within a room that are solid-walled and
bottomed areas for containing multiple cages housing infected animals.
The containment for these areas is equivalent to the Class I biological
safety cabinet. For testing purposes, they will be treated the same as a
Class I biological safety cabinet.
Appendix A to Part 627--References
Publications referenced in this part can be obtained from the
National Technical Information Services, U.S. Department of Commerce,
5285 Port Royal Road, Springfield, VA 22161.
Required Publications
AR 11-34
Army Respiratory Protection Program. (Cited in Secs. 627.31(h)(2)
and 627.31(h)(4).)
AR 40-5
Preventive Medicine. (Cited in Sec. 627.8.)
AR 40-10
Health Hazard Assessment Program in Support of the Army Materiel
Acquisition Decision Process. (Cited in Sec. 627.7(a)(8).)
AR 40-12
Medical and Agricultural Foreign and Domestic Quarantine Regulations
for Vessels, Aircraft, and Other Transports of the Armed Forces. (Cited
in Sec. 627.40(a).)
AR 40-66
Medical Records and Quality Assurance Administration. (Cited in
Sec. 627.9.)
AR 40-400
Patient Administration. (Cited in Sec. 627.8(e).)
AR 70-65
Management of Controlled Substances, Ethyl Alcohol, and Hazardous
Biological Substances in Army Research, Development, Test, and
Evaluation Facilities. (Cited in Secs. 627.36(a)(6) and 627.40(b).)
AR 385-10
Army Safety Program. (Cited in Secs. 627.6 and 627.31(h)(4).)
AR 385-69
Biological Defense Safety Program. (Cited in Secs. 627.6, 627.7(a),
627.7(a)(8), 627.7(d), 627.11(c), 627.18(a) and 627.18(f)(1).)
AR 740-32
Responsibilities for Technical Escort of Dangerous Materials. (Cited
in Sec. 627.39.)
Related Publications
A related publication is merely a source of additional information.
The user does not have to read it to understand this pamphlet.
AR 40-14
Control and Recording Procedures for Exposure to Ionizing Radiation
and Radioactive Materials.
ANSI Z86.1-1973
Breathing Air
ASHRAE Standard 62
Bacterial Toxins: A Table of Lethal Amounts, Gill, D.M.,
Microbiological Reviews, Volume 46, Number 1; March 1982, pages 86-94.
Biohazards Reference Manual
American Industrial Hygiene Association, 1985, Clinical Medicine
Branch, Division of Host Factors, Center for Infectious Disease, Centers
for Disease Control, Atlanta, GA 30333, telephone: (404) 639-3356,
Compressed Gas Association Pamphlet G-7.1
Grade D Breathing Air
Dangerous Goods Regulations, International Air Transport Association
(IATA), Publications Section, 2000 Peel Street, Montreal, Quebec, Canada
H3A 2R4, Tel (514) 844-6311. DHEW Pub. No. (NIH) 76-1165
Biological Safety Manual for Research Involving Oncogenic Viruses,
Executive Order 12196
Safety and Health Programs for Federal Employees, 26 February 1980
Guide for Adult Immunizations, Published by the American College of
Physicians, Guide for Transportation of Hazardous Materials, Vol. 4(1)
February 10, 1975. (Copies may be obtained from the Office of Research
Grants Inquiries, NIH, Department of Health and Human Services, 5333
Westbard Avenue, Bethesda, MD 20205.)
Guidelines for Laboratory Design, Health and Safety Considerations,
L. DiBerardinis, et al., John Wiley and Sons, 1987
Guidelines for Prevention of Herpesvirus Simiae (B Virus) Infection
in Monkey Handlers, Kaplan, J.E., et al., Mortality and Morbidity Weekly
Report, Volume 36, Number 41; October 23, 1987, pages 680-689.
HHS Publication No. (NIH) 88-8395, Biosafety in Microbiological and
Biomedical Laboratories
Industrial Ventilation, A Manual of Recommended Practice Published
by the American Conference of Governmental Industrial Hygienists.
[[Page 570]]
Laboratory Safety for Arboviruses and Certain Other Viruses of
Vertebrates, The American Journal of Tropical Medicine and Hygiene,
29:1359-1381, 1980.
NIH Guidelines for Research involving Recombinant DNA Molecules (51
FR 16958, May 7, 1986).
NIH publication 86-23, Guide for the Care and Use of Laboratory
Animals
NSF Standard 49, National Sanitation Foundation Standard
Number 49, Class II (Laminar Flow) Biohazard Cabinetry
Packaging and Shipping of Biological Materials at ATCC, The American
Type Culture Collection (ATCC). (Copies may be obtained from the ATCC,
12301 Parklawn Drive, Rockville, MD 20852. Telephone (301) 881-2600.)
Postal Bulletin No. 21246, International Mail-Hazardous Materials
Procedures for the Domestic Handling and Transport of Diagnostic
Specimens and Etiologic Agents, National Committee for Clinical
Laboratory Standards (NCCLS), (H5-A2), Second edition. Vol. 5, No. 1.
(Copies may be obtained from the NCCLS, 771 East Lancaster Avenue,
Villanova, PA 19085.)
Restricted Articles Tariff 6-D, Air Transport Association
Technical Instructions for the Safe Transport of Dangerous Goods by
Air, International Civil Aviation Organization (ICAO) Intereg Group,
5724 Pulaski Road, Chicago, IL 60646, Tel. (312) 478-0900.
The Centers for Disease Control, Office of Biosafety, 1600 Clifton
Road NE., Atlanta, Georgia 30333. Telephone (404) 639-3883, or FTS: 236-
3883.
9 CFR Parts 102 Through 104, 122
Animals and Animal products.
10 CFR Chapter 1
Nuclear Regulatory Commission.
21 CFR Parts 312, 600 Through 680
Food and drugs.
29 CFR Part 1910
Occupational Health and Safety Administration Safety and Health
Standards.
39 CFR Part 111
Postal Service.
40 CFR Parts 1500 Through 1508
Protection of environment.
42 CFR Parts 71 and 72
Public Health Service Foreign Quarantine Regulations.
49 CFR Parts 172 and 173
The Department of Transportation.
[[Page 571]]
Appendix B to Part 627--Resource List for Immunoprophylaxis of Personnel
at Risk
B-1. Recommendations for Immunoprophylaxis of Personnel at Risk
----------------------------------------------------------------------------------------------------------------
Description of disease Product Recommended for use in Source of product
----------------------------------------------------------------------------------------------------------------
Anthrax.......................... Inactivated vaccine. Personnel working regularly with USAMRIID.\1\
cultures, diagnostic materials,
or infected animals.
Botulism......................... Pentavalent toxoid Personnel working regularly with CDC.\3\
(A,B,C,D,E) cultures or toxin.
(IND).\2\.
Cholera.......................... Inactivated vaccine. Personnel working regularly with Commercially
large volumes or high available.
concentrations of infectious
materials.
Diphtheria Tetanus (Adult)....... Combined toxoid..... All laboratory and animal care Commercially
personnel irrespective of agents available.
handled.
Eastern equine encephalitis (EEE) Inactivated vaccine Personnel who work directly and USAMRIID.\1\
(IND) \2\. regularly with EEE in the
laboratory.
Hepatitis A...................... Immune Serum Animal care personnel working Commercially
Globulin [ISG directly with chimpanzees available.
(Human)]. naturally or experimentally
infected with Hepatitis A virus.
Hepatitis B...................... Serum-derived or Personnel working regularly with Commercially
recombinant vaccine. human blood and blood components. available.
Influenza........................ Inactivated vaccine. (Vaccines prepared from earlier Commercially
isolated strains may be of little available.
value in personnel working with
recent isolates from humans or
animals).
Japanese Encephalitis............ Inactivated vaccine Personnel who work directly and CDC.\3\
(IND) \2\. regularly with JE virus in the
laboratory.
Measles.......................... Live attenuated Measles-susceptible personnel Commercially
virus vaccine. working with the agent or available.
potentially infectious clinical
materials.
Meningococcal Meningitis......... Purified Personnel working regularly with Commercially
polysaccharide large volumes or high available.
vaccine. concentrations of infectious
materials (does not protect
against infection with group B
meningococcus).
Plague........................... Inactivated vaccine. Personnel working regularly with Commercially
cultures of Yersinia pestis or available.
infected rodents or fleas.
Poliomyelitis.................... Inactivated (IPV) Polio-susceptible personnel Commercially
and live attenuated working with the virus or available.
(OPV) vaccines. entering laboratories or animal
rooms where the virus is in use.
Pox viruses (Vaccinia, Cowpox, or Live (lyophilized) Personnel working with orthopox CDC.\3\
Monkey Pox viruses). vaccinia virus. viruses transmissible to humans,
with animals infected with these
agents, and persons entering
areas where these viruses are in
use.
Q Fever (Phase II) vaccine....... Inactivated (IND) Personnel who have no demonstrable USAMRIID.\1\
\2\. sensitivity to Q fever antigen
and who are at high risk of
exposure to infectious materials
or animals.
Rabies........................... Human diploid line Personnel working with all strains Commercially
cell inactivated of rabies virus, with infected available.
vaccine. animals, or persons entering
areas where these activities are
conducted.
Rift Valley Fever................ Inactivated virus All laboratory and animal care USAMRIID.\1\
vaccine (IND) \2\. personnel working with the agent
or infected animals and all
personnel entering laboratories
or animal rooms when the agent is
in use.
Rubella.......................... Live attenuated Rubella-susceptible personnel, Commercially
virus vaccine. especially women, working with available.
``wild'' strains or in areas
where these viruses are in use.
Tuberculosis..................... Live, attenuated BCG vaccine ordinarily is not used Commercially
(BCG) bacterial in laboratory personnel in the available.
vaccine. U.S.
Tularemia........................ Live attenuated Personnel working regularly with USAMRIID.\1\
bacterial vaccine cultures or infected animals or
(IND).\2\. persons entering areas where the
agent of infected animals are in
use.
Typhoid.......................... Inactivated vaccine. Personnel who have no demonstrated Commercially
sensitivity to the vaccine and available.
who work regularly with cultures.
Venezuelan equine (VEE) Live attenuated Personnel working with VEE and the USAMRIID.\1\
encephalitis. (TC83) viral Equine Cabassou, Everglades,
vaccine (IND).\2\. Mucambo, and Tonate viruses, or
who enter areas where these
viruses are in use.
[[Page 572]]
Western equine encephalitis (WEE) Inactivated vaccine Personnel who work directly and USAMRIID.\1\
(IND) \2\ with WEE regularly in the laboratory.
virus.
Yellow Fever..................... Live attenuated Personnel working with virulent Commercially
(17D) virus vaccine. and avirulent strains of Yellow available.
Fever virus.
----------------------------------------------------------------------------------------------------------------
\1\ For information, contact: United States Army Medical Materiel Development Activity, Fort Detrick, Frederick,
MD 21701, telephone: (301) 663-7661.
\2\ Investigational New Drug (IND).
\3\ Clinical Medicine Branch, Division of Host Factors, Center for Infectious Disease, Centers for Disease
Control, Atlanta, GA 30333, telephone: (404) 639-3356.
Source: Adapted from recommendations of the PHS Immunization Practices Advisory Committee and Biosafety in
Microbiological and Biomedical Laboratories.
[[Page 573]]
Appendix C to Part 627--Laboratory Safety Inspection Checklist
C-1. The checklist that follows is not an exhaustive list of the
items to consider when inspecting facilities where etiologic agents are
used. It does provide some basic guidelines to remind safety and
nonsafety professionals of the things that need to be considered in the
laboratories they manage. The checklist should be used as follows: All
area should be inspected using the general list in C-2. Certain items
are optional, such as radiation safety. If no radioactive material is
present in the room, then this would not be applicable. For BL-1
facilities the list in C-2 is adequate, while BL-2, BL-3, and BL-4
facilities must use the list in C-2 together with the appropriate list
in C-3 to C-5.
C-2. Basic checklist
(a) Housekeeping
(1) Is the room free of clutter?
(2) Are all aisles from the work areas to the available exits
maintained clear of obstructions?
(3) Are all safety equipment items unobstructed and ready for use?
(4) Is the room clean?
(b) Fire safety
(1) Is the fire extinguisher hung in its proper place, ready for
use, and unobstructed?
(2) Are there excess flammables located outside National Fire
Protection Association (NFPA) approved cabinetry?
(3) Are all Class IA flammables that are in breakable containers in
pint or smaller containers?
(4) Are all Class IB flammables that are in breakable containers in
liter or smaller containers?
(c) Chemical safety
(1) Are the chemicals stored with compatible materials?
(2) Have the chemical fume hoods been certified in the last 6
months?
(3) Are the eyewash and deluge shower unobstructed and ready for
use?
(4) Is the eyewash and deluge shower tested regularly to document
proper operation?
(5) Is the organic waste container maintained in a closed position?
(6) Are all reagents and solutions properly labeled?
(7) Is a spill kit within a reasonable distance from the work areas?
(8) Is appropriate protective clothing available for the chemical
hazards present?
(9) Is there a written hazard communication program?
(10) Have the personnel in the laboratory been trained in the
provisions and principles of the hazard communication program?
(11) Are MSDSs located where they are available to the laboratory
workers?
(12) Is there a written chemical hygiene plan?
(d) Radiation safety
(1) Are the radioactive materials stored double-contained?
(2) Is the containment for the radiation waste container adequate to
preclude the spread of radiation?
(3) Are all containers appropriately labeled with radiation labels?
(4) Are all entrances to the room appropriately labeled?
(e) Electrical safety
(1) Are excess extension cords being utilized?
(2) Are there any frayed cords in the room?
(3) Are there any cords on the floor across normal traffic patterns
in the room?
(f) General laboratory safety
(1) Are sharps discarded and destroyed in a safe manner?
(2) Are work surfaces decontaminated daily and after a spill?
(3) Is the appropriate attire worn by everyone in the room?
(4) Is there evidence that personnel eat, drink, smoke, or store
food, drinks, or tobacco in the room?
(5) Was mouth pipetting observed?
(6) Are all gas cylinders secured and are all cylinders not in use
capped?
(7) Are cylinders of oxidizers stored at least 20 feet from
cylinders of flammable gases in the same room?
(8) Are the contents of the cylinders clearly labeled?
(9) Are the cylinders transported on appropriate dollies or hand
trucks?
(10) Is there a written respiratory protection program where
respirators are used?
(g) Etiologic agents
(1) Are all containers of etiologic agents appropriately labeled?
(i) Are freezers, refrigerators, and similar storage units labeled
with the biohazard warning sign?
(ii) Are the storage and shipping containers adequate and properly
labeled?
(2) Have all personnel been adequately trained in general
microbiological techniques?
(3) Are laboratory doors kept closed when experiments are in
progress?
(4) Are all operations conducted over plastic-backed absorbent paper
or spill trays?
C-3. Biosafety level 2 supplemental checklist
(a) Are all floor drains filled with water or suitable disinfectant?
(b) Is the SOP for an etiologic agent spill signed by all personnel
who work with etiologic agents in the room?
(c) If biological safety cabinets are used, have they been certified
within the last year?
(d) Are the appropriate decontaminants available?
(e) Are all entrances to the laboratory posted with--
[[Page 574]]
(1) The appropriate special provisions for entry?
(2) The universal biohazard symbol?
(3) The name and telephone number of the laboratory director or
other responsible person?
(f) Is entry limited and restricted?
(g) Are gloves being worn when handling infected animals or
infectious or toxic materials?
(h) Is eye and respiratory protection being worn in rooms where
nonhuman primates are present?
(i) If materials are being transported off-site for decontamination,
is the containment adequate?
C-4. Biosafety level 3 supplemental checklist
(a) Is laboratory clothing decontaminated before being sent to the
laundry?
(b) Are all windows and penetrations through the walls and ceilings
sealed?
(c) If biological safety cabinets are used, have they been certified
within the last year?
(d) Are the appropriate decontaminants available?
(e) Are all entrances to the facility posted with--
(1) The appropriate special provisions for entry?
(2) The universal biohazard symbol?
(3) The name and telephone number of the laboratory director or
other responsible person?
(f) Is entry limited and restricted?
(g) Are gloves being worn when handling infected animals or
infectious or toxic materials?
(h) Is eye and respiratory protection being worn in rooms where
nonhuman primates are present?
(i) Do the monitors indicate that the room is under negative
pressure relative to all entrances?
(j) Are all vacuum lines protected with HEPA filters and liquid
disinfectant traps?
(k) Is the autoclave being properly maintained and certified?
(l) Is the foot, elbow, or automatic handwash sink operating
properly?
(m) Are all operations with etiologic agents being conducted inside
biological safety cabinets or other approved engineering controls?
(n) Are all infected animals housed using appropriate primary
containment systems?
(o) Do all personnel who enter rooms housing infected animals wear
appropriate respiratory protection?
(p) Do personnel who exit rooms having infected animals leave their
protective clothing in the animal and laboratory rooms?
(q) If available, has the UV pass box ouput been certified within
the last 3 months?
C-5. Biosafety level 4 supplemental inspection checklist
(a) Precautions for all areas.
(1) Are all penetrations through the walls and ceilings sealed?
(2) Are the appropriate decontaminants available and used properly?
(3) Are all entrances to the facility posted with--
(i) The appropriate special provisions for entry?
(ii) The universal biohazard symbol?
(iii) The name and telephone number of the laboratory director or
other responsible person?
(4) Is access to the laboratory controlled strictly and documented?
(5) Do the monitors indicate that the room is under negative
pressure relative to all entrances?
(6) Are all vacuum lines protected with HEPA filters and liquid
disinfectant traps?
(7) Is the autoclave being properly maintained and certified?
(8) Is the foot, elbow, or automatic handwash sink operating
properly?
(9) Do the self-closing doors to the facility operate properly?
(10) Do personnel completely exchange street clothing for laboratory
clothing before entry and shower upon exiting?
(11) Is the dunk tank disinfectant fresh and appropriate for the
agents in use?
(b) Suit areas.
(1) Are all operations with etiologic agents conducted in Class I or
II biological safety cabinets?
(2) Do the procedures in place ensure that, as much as possible, the
contamination remains inside the cabinets (such as ensuring that
everything removed from within the cabinets, such as gloves being worn,
instruments, glassware, or similar items, are decontaminated or properly
packaged first)?
(3) Are the Class I or II cabinets in the facility certified every 6
months?
(4) Does the suit decontamination shower have adequate appropriate
decontaminant available?
(5) Has the suit decontamination shower been used or tested in the
last month?
(6) Is the ventilated suit air supply and emergency air supply
adequate and working properly?
(7) Is the emergency alarm system working properly?
(8) Are all of the one-piece positive pressure suits available for
use in serviceable condition?
(9) Are infected animals housed in appropriate primary containment
systems?
(10) Is the static pressure in the suit area negative to all
surrounding areas?
(c) Nonsuit areas.
(1) Are all operations with etiologic agents conducted inside Class
III biological safety cabinets?
[[Page 575]]
(2) Were the Class III biological safety cabinets certified before
initiating the current operation?
(3) Are all infected animals housed in Class III cabinet containment
caging systems?
Appendix D to Part 627--Packaging and Labeling Requirements for Shipment
of Etiologic Agents
D-1. Packaging and Labeling of Etiologic Agents, from HHS publication
No. (NIH) 88-8395.
D-2. Guidelines for the Air Shipment of Diagnostic Specimens, from the
Air Transport Association of America, Cargo Services Division, 1709 New
York Ave., NW., Washington, DC 20006.
Appendix E to Part 627--Permits for Importation and Shipment of
Etiologic Agents
E-1. Permit Application to Import or Transport Agents or Vectors of
Human Disease. Department of Health, Education and Welfare, PHS, CDC,
Office of Biosafety, Atlanta, Georgia 30333.
E-2. Permit Application to Import Controlled Material; Import or
Transport Organisms or Vectors. U.S. Department of Agriculture, Animal
and Plant Health Inspection Service, Veterinary Services, Federal
Building, Hyattsville, Maryland 20782.
Appendix F to Part 627--Drawings, Biological Safety Cabinets
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Appendix G to Part 627--Glossary
Abbreviations
AIHA American Industrial Hygiene Association
AMC United States Army Materiel Command
ANSI American National Standards Institute
AR Army Regulation
ATCC American Type Culture Collection
ASHRAE American Society of Heating, Refrigerating, and
Air Condition Engineers, Inc.
BDP Biological Defense Program
BL biosafety level
CDC Centers for Disease Control
CFR Code of Federal Regulations
DA PAM Department of Army Pamphlet
DHEW Department of Health, Education, and Welfare
DOD Department of Defense
DOT Department of Transportation
DNA deoxyribonucleic acid
EPA Environmental Protection Agency
EtO ethylene oxide
FDA Food and Drug Administration
fpm feet per minute
HEPA high efficiency particulate air
[[Page 579]]
HHS Health and Human Services
IATA International Air Transport Association
IBC Institutional Biosafety Committee
ICAO International Civil Aviation Organization
lfpm linear feet per minute
LS large-scale
m meter
min minute
MSDS Material Safety Data Sheets
MSHA Mine Safety and Health Administration
NCCLS National Committee for Clinical Laboratory
Standards
NCI National Cancer Institute
NEPA National Environmental Policy Act
NFPA National Fire Protection Association
NIH National Institutes of Health
NIOSH National Institute for Occupational Safety and
Health
NRC Nuclear Regulatory Commission
NSF National Sanitation Foundation
OSHA Occupational Safety and Health Administration
pH the negative logarithm of hydrogen ion
concentration
PHS Public Health Service
PPE personal protective equipment
ppm parts per million
psi pounds per square inch
RCRA-Listed Resource Conservation Recovery Act of 1976
Listed Hazardous Waste
RDTE research, development, test, and evaluation
RPO Radiation Protection Officer
SALS Subcommittee on Arbovirus Laboratory Safety
SAR supplied-air respirator
SCBA self-contained breathing apparatus
SOP Standing Operating Procedure
TD to deliver
TLV threshold limit value
USDA United States Department of Agriculture
UV ultraviolet
Terms
Approved respiratory protection
Equipment which is tested and listed as satisfactory according to
standards established by a competent authority (such as NIOSH, Mine
Safety and Health Administration (MSHA), or host country agency) to
provide respiratory protection against the particular hazard for which
it is designed. For military agent protection, DA and Department of
Defense (DOD) are the approval authorities. (Approval authority may be
specified by law.)
Biocontainment area
An area which meets the requirements for a BL-3 or BL-4 facility.
The area may be an entire building or a single room within a building.
See subpart G for details.
Biological Safety Cabinets
Engineering controls designed to enable laboratory workers to handle
infectious etiologic agents and to provide primary containment of any
resultant aerosol. There are three major classes of cabinets (I, II, and
III) and several subclasses of class II cabinets. Each type of cabinet
provides a different degree of protection to personnel and to the
products handled inside them. The various classes of cabinets are
described in detail in subpart H.
Biosafety Level 1
The facilities, equipment, and procedures suitable for work
involving agents of no known or of minimal potential hazard to
laboratory personnel and the environment.
Biosafety Level 2
The facilities, equipment, and procedures applicable to clinical,
diagnostic, or teaching laboratories, and suitable for work involving
indigenous agents of moderate potential hazard to personnel and the
environment. It differs from BL-1 in that (1) laboratory personnel have
specific training in handling pathogenic agents, (2) the laboratory is
directed by scientists with experience in the handling of specific
agents, (3) access to the laboratory is limited when work is being
conducted, and (4) certain procedures in which infectious aerosols could
be created are conducted in biological safety cabinets or other physical
containment equipment.
Biosafety Level 3
The facilities, equipment, and procedures applicable to clinical,
diagnostic, research, or production facilities in which work is
performed with indigenous or exotic agents where potential exists for
infection by aerosol, and the disease may have serious or lethal
consequences. It differs from BL-2 in that (1) more extensive training
in handling pathogenic and potentially lethal agents is necessary for
laboratory personnel; (2) all procedures involving the manipulation of
infectious material are conducted within biological safety cabinets,
other physical containment devices, or by personnel wearing appropriate
personal protective clothing and devices; (3) the laboratory has special
engineering and design features, including access zones, sealed
penetrations, and directional airflow; and (4) any modification of BL-3
recommendations must be made only by the commander.
Biosafety Level 4
The facilities, equipment, and procedures required for work with
dangerous and exotic agents which pose a high individual risk of life-
threatening disease. It differs from BL-3 in that (1) members of the
laboratory staff have specific and thorough training in handling
extremely hazardous infectious agents; (2) laboratory personnel
understand the primary and secondary containment functions of the
standard and special practices, containment equipment, and laboratory
design characteristics; (3) access to the laboratory is strictly
controlled by the institute director; (4) the facility is either in a
separate
[[Page 580]]
building or in a controlled area within a building, completely isolated
from all other areas of the building; (5) a specific facility operations
manual is prepared or adopted; (6) within work areas of the facility,
all activities are confined to Class III biological safety cabinets or
Class I or Class II biological safety cabinets used in conjunction with
one-piece positive pressure personnel suits ventilated by a life support
system; and (7) the maximum containment laboratory has special
engineering and design features to prevent microorganisms from being
disseminated to the environment.
Building
A structure that contains the requisite components necessary to
support a facility that is designed according to the required biosafety
level. The building can contain one or more facilities conforming to one
or more biosafety level.
Confirmed Exposure
Any mishap with a BDP agent in which there was direct evidence of an
actual exposure such as a measurable rise in antibody titer to the agent
or a confirmed diagnosis of intoxication or disease.
Etiologic Agents
Any viable microorganism, or its toxin which causes or may cause
human disease, including those agents listed in 42 CFR 72.3 of the
Department of Health and Human Services regulations, and any agent of
biological origin that poses a degree of hazard similar to those agents.
Facility
An area within a building that provides appropriate protective
barriers for persons working in the facility and the environment
external to the facility, and outside of the building.
HEPA Filter
A filter which removes particulate matter down to submicron sized
particles from the air passed through it with a minimum efficiency of
99.97 percent. While the filters remove particulate matter with great
efficiency, vapors and gases (for example, from volatile chemicals) are
passed through without restriction. HEPA filters are used as the primary
means of removing infectious agents from air exhausted from engineering
controls and facilities.
Human Lethal Dose
The estimated quantity of a toxin that is a minimum lethal dose for
a 70 kilogram individual based upon published data or upon estimates
extrapolated from animal toxicity data.
Commander or Institute Director
The commander or institute director of an Army activity conducting
RDTE with BDP etiologic agents, or the equivalent, at a research
organization under contract to the BDP.
Institution
An organization such as an Army RDTE activity (institute, agency,
center, and so forth) or a contract organization such as a school of
medicine, or research institute that conducts RDTE with BDP etiologic
agents.
Laboratory
An individual room or rooms within a facility that provide space in
which work with etiologic agents can be performed. It contains all of
the appropriate engineering features and equipment required at a given
biosafety level to protect personnel working in it and the environment
external to the facility.
Large-Scale Operations
Research or production involving viable etiologic agents in
quantities greater than 10 liters of culture.
Maximum Containment Area
An area which meets the requirements for a BL-4 facility. The area
may be an entire building or a single room within the building. See
chapter 7 for details.
Molded Masks
Formed masks that fit snugly around the mouth and nose and are
designed to protect against a nontoxic nuisance level of dusts and
powders. These do not require approval by NIOSH or MSHA. Masks made of
gauze do not qualify.
Potential Accidental Exposure
Any accident in which there was reason to believe that anyone
working with a BDP agent may have been exposed to that agent, yet no
measurable rise in antibody titer or diagnosis of intoxication or
disease was made. However, the high probability existed for introduction
of an agent through mucous membranes, respiratory tract, broken skin, or
the circulatory system as a direct result of the accident, injury, or
incident.
Resource Conservation Recovery Act of 1976 Listed Hazardous Waste
The waste materials listed by the Environmental Protection Agency
under authority of the RCRA for which the agency regulates disposal. A
description and listing of these wastes is located in 40 CFR part 261.
[[Page 581]]
Suite
An area consisting of more than one room, designed to be a
functional unit in which entire operations can be facilitated. Suites
may contain a combination of laboratories or animal holding rooms and
associated support areas within a facility that are designed to conform
to a particular biosafety level. There may be one or more suites within
a facility.
Toxin
Toxic material of etiologic origin that has been isolated from the
parent organism.\1\
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\1\ The publication ``Bacterial Toxins: a Table of Lethal Amounts,''
(Gill, D.M. (1982) Microbiological Reviews, 46:86-94) contains a useful
table of mammalian toxicities of numerous toxins.