[Federal Register Volume 59, Number 54 (Monday, March 21, 1994)]
[Unknown Section]
[Page 0]
From the Federal Register Online via the Government Publishing Office [www.gpo.gov]
[FR Doc No: 94-6550]


[[Page Unknown]]

[Federal Register: March 21, 1994]


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DEPARTMENT OF AGRICULTURE
9 CFR Part 113

[Docket No. 93-071-1]

 

Viruses, Serums, Toxins, and Analogous Products; Detection of 
Extraneous Agents by the Fluorescent Antibody Technique

AGENCY: Animal and Plant Health Inspection Service, USDA.

ACTION: Proposed rule.

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SUMMARY: We are proposing to amend the regulations concerning testing 
by the fluorescent antibody technique for extraneous agents (viruses) 
in cells, of animal origin that are used in the manufacture of 
veterinary biologics. The proposed amendment would allow the use of 
alternative fluorescent-antibody conjugated antibodies; revised the 
list of extraneous agents to be tested for; and include extraneous 
agents for which equine cells are to be tested. The proposed amendment 
is necessary to update the requirements related to the testing for 
extraneous viruses.

DATES: Consideration will be given only to comments received on or 
before April 20, 1994.

ADDRESSES: Please send an original and three copies of your comments to 
Chief, Regulatory Analysis and Development, PPD, APHIS, USDA, room 804, 
Federal Building, 6505 Belcrest Road, Hyattsville, MD 20782. Please 
state that your comments refer to Docket No. 93-071-1. Comments 
received may be inspected at USDA, room 1141, South Building, 14th 
Street and Independence Avenue SW., Washington, DC, between 8 a.m. and 
4:30 p.m., Monday through Friday, except holidays. Persons wishing to 
inspect comments are requested to call ahead on (202) 690-2817 to 
facilitate entry into the comment reading room.

FOR FURTHER INFORMATION CONTACT:
Dr. David A. Espeseth, Deputy Director, Veterinary Biologics, BBEP, 
APHIS, USDA, room 838, Federal Building, 6505 Belcrest Road, 
Hyattsville, MD 20782, (301) 436-8245.

SUPPLEMENTARY INFORMATION:

Background

    In accordance with the regulations contained in 9 CFR part 113, 
Standard Requirements are prescribed for the preparation of veterinary 
biological products. A Standard Requirement consists of specifications, 
procedures, and test methods which define the standards of purity, 
safety, potency, and efficacy for a given type of veterinary biological 
product. Microorganisms, animal cells, and ingredients of animal origin 
used in production are required by regulations in part 113 to be tested 
for extraneous viruses. In part, this involves testing for the presence 
of extraneous viruses by the fluorescent antibody technique described 
in Sec. 113.47. When the current standard requirement was established, 
fluorescent antibodies were constructed by conjugating antibodies to 
one of the fluorochromes, fluorescein. Fluorochromes are any of a 
variety of chemicals used in cytochemistry to produce a secondary 
fluorescence in the specimen. In the intervening years, additional 
fluorochromes have been developed for use as cytochemical markers or 
stains.
    Standard Requirements included in the regulations specify that 
cells, Master Seed virus, and most ingredients of animal origin used in 
the production of biological products be tested for contaminating 
bacteria, fungi, mycoplasma, cytopathogenic organisms, viruses, 
hemadsorbing agents, and extraneous agents (viruses) detectable by the 
fluorescent antibody technique. The presence of specific fluorescence 
associated with the use of certain antibodies, in comparison with the 
appropriate controls, is an indication of the presence of the 
contaminating antigen or extraneous virus against which the antibody 
was made.
    Current Sec. 113.47 lists the types of extraneous viruses against 
which fluorescein-conjugated antibodies are to be used in testing cells 
from certain species of animals. New viruses have since been identified 
as animal pathogens. No viruses which are disease agents of horses are 
included in the current Sec. 113.47. As new knowledge has developed, 
testing for these agents has been necessary.

Changes and Clarifications

    The title of Sec. 113.47 refers to the detection of extraneous 
agents. Since the section is specific to viruses, the title would be 
amended accordingly.
    The current regulations in Sec. 113.47 limit the stain used in the 
fluorescent antibody test to fluorescein (a fluorochrome). Other 
fluorochromes, when conjugated to antibodies, may be expected to 
perform as well as fluorescein in the test for extraneous viruses. This 
proposed amendment would allow the use of any fluorochrome-conjugated 
antibody in such tests. The term ``fluorescein-conjugated antibody'' 
would be replaced with ``fluorochrome-conjugated antibody'' everywhere 
it appears in Sec. 113.47 and in Sec. 113.52(b)(2) (i) and (ii).
    The current regulation in Sec. 113.47(a) provides for staining cell 
monolayers for parvovirus as early as 4 days after subculturing. Other 
viruses may also be detected by processing at less than 7 days. 
Therefore, the specific exemption for parvovirus has been deleted in 
Sec. 113.47(a), and the provision in Sec. 113.47(a)(3) for fixing 
control monolayers at less than 7 days has been modified to require 
that a test monolayer also be fixed (processed so as to arrest growth 
and assure attachment of the monolayer to the surface of the vessel in 
which they are grown) at the same time. In addition, another monolayer 
of the material under test and the negative control monolayer must be 
held for at least 7 days before fixing. Further clarification is made 
to Sec. 113.47(a)(3) that all monolayers would be stained concurrently, 
regardless of time of fixing, at least 7 days after subculture.
    The current regulations in Sec. 113.47 lists the specific 
extraneous viruses against which antibodies are to be used in the 
testing of certain types of cells. The cells with which the fluorescent 
antibody test is to be used are specified in Secs. 113.51, 113.52, 
113.53, or 113.55. In the proposed amendment to Sec. 113.47, Vero cells 
would not be listed as one of the specific cell types since they are 
specified in those sections as listed above. We have also revised the 
list of cell types to be tested for extraneous viruses to include 
equine cells. Those using other cells for the production of biologics 
may also be required to test for specified viruses before such use is 
approved.
    Proposed amendments to Sec. 113.47(b) pertain to the extraneous 
viruses associated with specific cells that would be tested for using 
specific fluorochrome-conjugated antibodies. All cells (including 
caprine and equine would be tested for bovine virus diarrhea virus, 
reovirus, and rabies virus. In addition, bluetongue virus would be 
added to the list of extraneous agents for testing bovine cells, canine 
coronavirus would be listed for testing canine cells, feline infectious 
peritonitis virus would be specified in place of coronavirus for 
testing feline cells, and porcine hemagglutinating encephalitis virus 
would be added for testing porcine cells. Equine cells would be tested 
with fluorochrome-conjugated antibodies to equine herpesvirus and to 
equine viral arteritis virus.
    In addition, non-substantive minor editorial changes to the 
regulations are made for clarity.

Executive Order 12866 and Regulatory Flexibility Act

    This rule has been determined to be not significant for purpose of 
Executive Order 12866 and therefore has not been reviewed by OMB.
    These proposed amendments should not have a significant economic 
impact on manufacturers since it would broaden the range of 
fluorochrome-labeled antibodies that may be used in conducting the 
fluorescent antibody test and would revise the list of extraneous 
agents for which various cell types are to be tested with the 
fluorescent antibody technique. The proposed amendments would thus 
remove outdated requirements and provide flexibility in the types of 
antibody that may be used in tests for extraneous agents.
    Under these circumstances, the Administrator of the Animal and 
Plant Health Inspection Service has determined that this action would 
not have a significant economic impact on a substantial number of small 
entities.

Executive Order 12372

    This program/activity is listed in the Catalog of Federal Domestic 
Assistance under No. 10.025 and is subject to Executive Order 12372, 
which requires intergovernmental consultation with State and local 
officials. (See 7 CFR part 3015, subpart V.)

Executive Order 12778

    This proposed rule has been reviewed under Executive Order 12778, 
Civil Justice Reform. If this proposed rule is adopted: (1) All State 
and local laws and regulations that are in conflict with this rule will 
be preempted; (2) no retroactive effect will be given to this rule; and 
(3) administrative proceedings will not be required before parties may 
file suit in court challenging this rule.

Paperwork Reduction Act

    This proposed rule contains no information collection or 
recordkeeping requirements under the Paperwork Act of 1980 (44 U.S.C. 
3501 et seq.)

List of Subjects in 9 CFR Part 113

    Animal biologics, Exports, Imports, Reporting and recordkeeping 
requirements.

    Accordingly, 9 CFR part 113 would be amended as follows:

PART 113--STANDARD REQUIREMENTS

    1. The authority citation for part 113 would continue to read as 
follows:

    Authority: 21 U.S.C. 151-159; 7 CFR 2.17, 2.51, and 371.2(d).

    2. Section 113.47 would be revised to read as follows:


Sec. 113.47  Detection of extraneous viruses by the fluorescent 
antibody technique.

    The test for detection of extraneous viruses by the fluorescent 
antibody technique provided in this section shall be conducted when 
prescribed in an applicable Standard Requirement or in a filed Outline 
of Production for a product.
    (a) Monolayer cultures of cells (monolayers), at least 7 days after 
the last subculturing, shall be processed and stained with the 
appropriate antiviral fluorochorome-conjugated antibody as specified in 
paragraph (b) of this action.
    (1) Three groups of one or more monolayers shall be required for 
each specific virus prescribed in paragraph (b) of this section.
    (i) At the time of the last subculturing, one group of test 
monolayers shall be inoculated with approximately 100-300 FAID50 
of the specific virus being tested for as positive controls.
    (ii) One group of monolayers shall be the ``material under test.''
    (iii) One group of monolayers, that are of the same type of cells 
as the test monolayers and that have been tested as prescribed in 
Secs. 113.51 or 113.52 (whichever is applicable), shall be prepared as 
negative controls.
    (2) Each group of monolayers shall have a total area of at least 6 
cm2.
    (3) Positive control monolayers may be fixed (processed so as to 
arrest growth and assure attachment of the monolayer to the surface of 
the vessel in which they are grown) before 7 days after subculturing if 
fluorescence is enhanced by doing so, Provided, That a monolayer of the 
material under test is also fixed at the same time as the positive 
control and a monolayer of the material under test is also fixed at 
least seven days after subculturing. Monolayers that are fixed before 7 
days after subculturing shall be stained at the same time as the test 
monolayers and negative controls fixed at least 7 days after 
subculturing.
    (b) The antiviral fluorochrome-conjugated antibodies to be used 
shall depend on the type of cells required to be tested for extraneous 
viruses as specified in an applicable Standard Requirement or in a 
filed Outline of Production. Antiviral fluorochrome-conjugated 
antibodies specific for the extraneous viruses shall be applied to each 
respective type of cell in accordance with the following list. Under 
certain circumstances, additional tests may need to be conducted, as 
determined by the Administrator. When a specific antiviral 
fluorochrome-conjugated antibody is used in testing for the listed 
extraneous viruses specified in more than one cell type, it need only 
be applied to the most susceptible cell type.

(1) All cells shall be tested for:
    (i) Bovine virus diarrhea virus;
    (ii) Reovirus; and
    (iii) Rabies virus.
(2) Bovine, caprine, and ovine cells shall, in addition, be tested for:
    (i) Bluetongue virus;
    (ii) Bovine adenoviruses;
    (iii) Bovine parvovirus; and
    (iv) Bovine respiratory syncytial virus.
(3) Canine calles shall, in addition, be tested for:
    (i) Canine coronavirus;
    (ii) Canine distemper virus; and
    (iii) Canine parvovirus.
(4) Equine cells shall, in addition, be tested for:
    (i) Equine herpesvirus; and
    (ii) Equine viral arteritis virus.
(5) Feline cells shall, in addition, be tested for:
    (i) Feline infectious peritonitis virus; and
    (ii) Feline panleukopenia virus.
(6) Porcine cells shall, in addition, be tested for:
    (i) Porcine adenovirus;
    (ii) Porcine parvovirus;
    (iii) Transmissible gastroenteritis virus; and
    (iv) Porcine hemagglutinating encephalitis virus.
(7) Firms that do not have rabies virus on premises either for research 
or production purposes are exempt from having to produce positive 
rabies virus control monolayers. Fixed positive rabies virus control 
monolayers will be provided by the National Veterinary Services 
Laboratories.

    (c) After staining, each group of monolayers shall be examined for 
the presence of specific fluorescence attributable to the presence of 
extraneous viruses.
    (1) If the material under test shows any evidence of specific viral 
fluorescence, it is unsatisfactory and may not be used; Provided, That, 
if specific fluorescence attributable to the virus being tested for is 
absent in the positive control monolayers, the test is inconclusive and 
may be repeated.
    (2) If the fluorescence of the monolayers inoculated with the 
specific virus as positive controls is equivocal, or if the negative 
monolayers show equivocal fluorescence indicating possible viral 
contamination, or both, the test shall be declared inconclusive, and 
may be repeated; Provided, That, if the test is not repeated, the 
material under test shall be regarded as unsatisfactory for use in the 
production of biologics.
    3. Section 113.52, paragraph (b)(2)(i) and (ii), would be revised 
to read as follows:


Sec. 113.52  requirements for cell lines used for production of 
biologics.

* * * * *
    (b) * * *
    (2) * * *
    (i) At least two monolayers shall be stained with an antispecies 
fluorochrome-conjugated antibody unrelated to the species of origin of 
the MCS.
    (ii) At least two monolayers shall be stained with an antispecies 
fluorochrome-conjugated antibody specific to the species of origin of 
the MCS.
* * * * *


Secs. 113.51, 113.52, and 113.53  [Amended]

    4. In the following places, the word ``agents'' would be removed 
and the word ``viruses'' added in its place:
    a. Section 113.51, paragraph (c)(3)(ii).
    b. Section 113.52, paragraph (f)(4)(ii).
    c. Section 113.53, paragraph (c)(6)(ii).

    Done in Washington, DC, this 15th day of March 1994.
Patricia Jensen,
Acting Assistant Secretary, Marketing and Inspection Services.
[FR Doc. 94-6550 Filed 3-18-94; 8:45 am]
BILLING CODE 3410-34-M