[Title 21 CFR 620]
[Code of Federal Regulations (annual edition) - April 1, 1996 Edition]
[Title 21 - FOOD AND DRUGS]
[Chapter I - FOOD AND DRUG ADMINISTRATION,]
[Subchapter F - BIOLOGICS]
[Part 620 - ADDITIONAL STANDARDS FOR BACTERIAL PRODUCTS]
[From the U.S. Government Publishing Office]
21
FOOD AND DRUGS
7
1996-04-01
1996-04-01
false
ADDITIONAL STANDARDS FOR BACTERIAL PRODUCTS
620
PART 620
FOOD AND DRUGS
FOOD AND DRUG ADMINISTRATION,
BIOLOGICS
PART 620--ADDITIONAL STANDARDS FOR BACTERIAL PRODUCTS--Table of Contents
Subpart A--Pertussis Vaccine
Sec.
620.1 Pertussis Vaccine.
620.2 Production.
620.3 U.S. Standard preparations.
620.4 Potency test.
620.5 Mouse toxicity test.
[[Page 71]]
620.6 General requirements.
Subpart B--Typhoid Vaccine
620.10 Typhoid Vaccine.
620.11 Production.
620.12 U.S. Standard preparations.
620.13 Potency test.
620.14 General requirements.
Subpart C--Anthrax Vaccine Adsorbed
620.20 Anthrax Vaccine Adsorbed.
620.21 Production.
620.22 U.S. Reference preparation.
620.23 Potency test.
620.24 General requirements.
Subpart D--Cholera Vaccine
620.30 Cholera Vaccine.
620.31 Production.
620.32 U.S. Standard preparations.
620.33 Potency tests.
620.34 Mouse toxicity test.
620.35 General requirements.
Subpart E--Bacillus of Calmette and Guerin (BCG) Vaccine
620.40 BCG Vaccine.
620.41 Establishment and personnel requirements.
620.42 Production.
620.43 Reference BCG Vaccine.
620.44 Potency tests.
620.45 Test for freedom from virulent mycobacteria.
620.46 General requirements.
620.47 Labeling.
620.48 Samples; protocols; official release.
Authority: Secs. 201, 501, 502, 503, 505, 510, 701 of the Federal
Food, Drug, and Cosmetic Act (21 U.S.C. 321, 351, 352, 353, 355, 360,
371); secs. 215, 351, 352, 353, 361 of the Public Health Service Act (42
U.S.C. 216, 262, 263, 263a, 264).
Source: 38 FR 32064, Nov. 20, 1973, unless otherwise noted.
Cross References: For U.S. Customs Service regulations relating to
viruses, serums, and toxins, see 19 CFR 12.21--12.23. For U.S. Postal
Service regulations relating to the admissibility to the United States
mails see parts 124 and 125 of the Domestic Mail Manual, that is
incorporated by reference in 39 CFR part 111.
Subpart A--Pertussis Vaccine
Sec. 620.1 Pertussis Vaccine.
The proper name of this product shall be ``Pertussis Vaccine'',
which shall be an aqueous preparation of killed whole Bordetella
pertussis bacteria. The vaccine may be precipitated or adsorbed and may
be combined with other antigens.
[56 FR 63410, Dec. 4, 1991]
Sec. 620.2 Production.
(a) Propagation of bacteria. Human blood shall not be used in
culture medium for propagating bacteria either for seed or for vaccine.
The culture medium for propagating bacteria for vaccine shall not
contain ingredients known to be capable of producing allergenic effects
in human subjects, except blood or blood products from lower animals
other than the horse. When blood or a blood product is used, it shall be
removed by washing the harvested bacteria. The bacterial concentrate
shall be free of extraneous bacteria, fungi, and yeasts, as demonstrated
by microscopic examination and cultural methods.
(b) Bacterial content. (1) The opacity of the bacterial concentrate
shall be determined in terms of the U.S. Opacity Standard not later than
2 weeks after the harvest of the bacteria and before any treatment
capable of altering the opacity of the bacterial concentrate.
(2) The total immunizing dose of a vaccine prepared with whole
bacteria shall contain (i) in the case of nonadsorbed vaccine no more
bacteria than the equivalent of 60 opacity units and (ii) in the case of
adsorbed vaccine no more than the equivalent of 48 opacity units.
(c) Detoxification. After removing a sample for purity testing, the
bacteria shall be killed and detoxified either (1) by heating, (2) by
addition of a chemical agent and appropriate aging, or (3) by any
combination of the stated procedures. The procedure used shall be one
that has been shown to have no adverse effect on required safety,
purity, and potency.
(d) Preservative. The vaccine shall contain a preservative.
Sec. 620.3 U.S. Standard preparations.
(a) The U.S. Standard Pertussis Vaccine shall be used for
determining the potency of Pertussis Vaccine.
(b) The U.S. Opacity Standard shall be used in estimating the
bacterial content of the vaccine and of the challenge culture.
[[Page 72]]
Sec. 620.4 Potency test.
The number of protective units of the total human immunizing dose
shall be estimated for each lot of vaccine from the results of
simultaneous intracerebral mouse protection tests of the vaccine under
test and the U.S. Standard Pertussis Vaccine. The potency test shall be
performed as follows:
(a) Mice. Healthy mice shall be used, all from a single strain and
of the same sex, or an equal number of each sex in each group, with
individual weight varying no more than 4 grams in a single test. In no
event shall any of the mice weigh less than 10 grams or more than 20
grams. A system of randomization shall be used to distribute the mice
into the groups, with respect to shelf position and to determine the
order of challenge. There shall be at least 3 groups consisting of no
less than 16 mice each, for each vaccine. In addition, there shall be at
least 4 groups consisting of no less than 10 mice each, for control
purposes: one group for the challenge dose and 3 groups for titrating
the virulence of the challenge dose.
(b) Vaccination. (1) Five-fold serial dilutions of the vaccine to be
tested and of the standard vaccine shall be made in 0.85 percent sodium
chloride solution. The dilutions of the vaccine under test shall have
the same protective unitage, based on an estimate of 12 units per total
human immunizing dose, as the unitage of the corresponding dilution of
the standard vaccine. Each mouse in each group for vaccination shall be
injected intraperitoneally with 0.5 ml. of the appropriate dilution.
(2) The interval between vaccination and challenge shall be 14 to 17
days. At least 87.5 percent of the mice in each group shall survive the
period between vaccination and challenge and each mouse challenged shall
appear healthy.
(c) The challenge. (1) The challenge culture of Bordetella pertussis
for each test shall be taken from a batch of cultures which have been
maintained by a method, such as freeze-drying, that retains constancy of
virulence.
(2) The challenge and virulence titration doses shall be prepared as
follows: The bacteria shall be harvested from a 20 to 24 hour culture
grown on Bordet-Gengou medium seeded from a rapidly growing culture less
than 48 hours old and uniformly suspended in a solution containing 1.0
percent casein peptone and about 0.6 percent sodium chloride at pH
7.1plus-minus0.1. The suspension, freed from agar particles and
clumps of bacteria, and adjusted to an opacity of 10 units, shall be
diluted in the solution used for suspending the bacteria, to provide in
a volume of 0.03 ml. (i) a challenge dose of 0.0001 opacity units
(1:3000) and (ii) virulence titration doses of \1/50\, \1/250\ and \1/
1250\ respectively of the challenge dose.
(3) Each vaccinated mouse shall be injected intracerebrally with the
challenge dose. The four groups of control mice shall be injected
intracerebrally with the challenge dose and its three dilutions,
respectively. The challenge-dose control mice shall be injected last.
The interval between the removal of the bacteria from the culture medium
and the injection of the last mouse shall not exceed 2\1/2\ hours.
(d) Recording the results. The mice shall be observed for 14 days.
Mice dying within 72 hours after challenge shall be excluded from the
test. Records shall be maintained of the number of mice that die after
72 hours and of the number of mice showing both paralysis and
enlargement of the head at the end of 14 days. All mice that show both
paralysis and enlargement of the head shall be considered as deaths for
the purposes of determining the ED50.
(e) Validity of the test. The test shall be valid provided (1) the
ED50 of the vaccine under test and the standard vaccine is between
the largest and smallest vaccinating doses; (2) the limits of one
standard deviation of each ED50 fall within the range of 64 percent
to 156 percent; (3) the protective response is graded in relation to the
vaccinating doses; (4) the dose-response curves of the vaccine under
test and the standard vaccine are parallel; (5) the challenge dose
contains approximately 200 LD50; (6) the LD50 contains no more
than 300 colony forming units; and (7) the \1/1250\ dilution of the
challenge dose contains no less than 10 and no more than 50 colony
forming units.
(f) Estimate of the potency. The ED50 of each vaccine shall be
calculated by a
[[Page 73]]
method that provides an estimate of the standard deviation. The
protective unit value per total human immunizing dose of the vaccine
under test shall be calculated in terms of the unit value of the
standard vaccine.
(g) Potency requirements. The vaccine shall have a potency of 12
units per total human immunizing dose based upon either a single test
estimate of no less than 8 units or a two-, three- or four-test
geometric mean estimate of no less than 9.6, 10.8, or 12 units,
respectively, except that for the vaccine in a multiple antigen product
containing Poliovirus Vaccine Inactivated, the estimate shall be no less
than 14 units. In no event shall the estimate be more than 36 units.
(h) Test design variation. Variations in the design of the potency
test may be permitted providing the results are demonstrated to be of
equal or greater precision.
[38 FR 32064, Nov. 20, 1973, as amended at 50 FR 4137, Jan. 29, 1985]
Sec. 620.5 Mouse toxicity test.
The final vaccine shall be demonstrated to be free from toxicity by
the following test:
A group of no less than 10 mice, each mouse weighing 14 to 16 grams,
shall have free access to food and water for no less than 2 hours before
injection. The group weight of the mice shall be determined immediately
prior to injection. Each mouse shall be injected intraperitoneally with
a test dose of one-half of the largest recommended single human dose of
the final vaccine in a volume of no less than 0.5 ml. nor more than 0.75
ml. The group weight of the mice shall be determined at the end of 72
hours and at the end of 7 days after injection. At the end of 72 hours
the average weight per mouse may be no less than the average weight per
mouse immediately preceding the injection; at the end of 7 days the
average weight gain per mouse may be no less than 3.0 grams; and at the
end of 7 days there may be vaccine-related deaths of no more than 5
percent of the total number of mice in all the toxicity tests performed.
Sec. 620.6 General requirements.
(a) Safety. Each lot of product containing Pertussis Vaccine shall
be tested for safety by the procedures prescribed in Sec. 610.11 of this
chapter except that the test shall consist of the intraperitoneal
injection of no less than one-half of the recommended largest individual
human dose into each of the mice, and either the intraperitoneal
injection of no less than three times the recommended largest individual
human dose, or the subcutaneous injection of 5.0 milliliters into each
of the guinea pigs.
(b) Dose. These additional standards are based on a single injection
of 0.5 ml., 1.0 ml., or 1.5 ml., and a total human immunizing dose of
three single injections of a nonadsorbed vaccine, and two or three
single injections of an adsorbed vaccine.
(c) Product characteristics. Recommendations shall be made through
appropriate labeling that the product after issue should not be frozen
and should be well shaken immediately prior to use.
(d) Labeling. In addition to the items required by other applicable
labeling provisions of this part, the package label shall give the
following information:
(1) For a vaccine containing a precipitant or an adsorbent, the word
``Adsorbed'' shall follow the proper name in the same style of type and
prominence as the proper name.
(2) The total immunizing dose contains 12 units of pertussis
vaccine.
(e) Multiple antigen products. The Pertussis Vaccine components of
multiple antigen products shall be manufactured pursuant to these
additional standards, except that the mouse toxicity test (Sec. 620.5)
and the potency test (Sec. 620.4) shall be performed on the multiple
antigen product.
(f) Adsorbed vaccines. Only aluminum compound reagents shall be
introduced into the product to cause precipitation or adsorption of
either Pertussis Vaccine or other antigens incorporated with Pertussis
Vaccine.
(g) Freezing prohibition. Pertussis Vaccine and multiple antigen
products of which Pertussis Vaccine is a component shall not be frozen
at any time during storage.
[[Page 74]]
(h) Samples and protocols. For each lot of vaccine, the following
material shall be submitted to the Director, Center for Biologics
Evaluation and Research, Food and Drug Administration, 8800 Rockville
Pike, Bethesda, MD 20892.
(1) A sample of no less than 20 milliliters of the final product for
pertussis vaccine testing.
(2) Protocols showing summaries of the manufacturing processes and
the results of all mouse toxicity (Sec. 620.5) and potency (Sec. 620.4)
tests performed.
[38 FR 32064, Nov. 20, 1973, as amended at 41 FR 35480, Aug. 23, 1976;
48 FR 13025, Mar. 29, 1983; 49 FR 23834, June 8, 1984; 51 FR 15610, Apr.
25, 1986; 55 FR 11013, Mar. 26, 1990]
Subpart B--Typhoid Vaccine
Sec. 620.10 Typhoid Vaccine.
The proper name of this product shall be Typhoid Vaccine which shall
be an aqueous or dried preparation of killed Salmonella typhi bacteria.
[48 FR 7167, Feb. 18, 1983]
Sec. 620.11 Production.
(a) Strain of bacteria. (1) Strain Ty 2 of Salmonella typhi shall be
used in the manufacture of Typhoid Vaccine.
(2) The antigenic integrity of the Ty 2 strain shall be verified by
an appropriate serological procedure.
(b) Propagation of bacteria. The culture medium for propagation of
S. typhi shall not contain ingredients known to be capable of producing
allergenic effects in human subjects. The harvested bacteria shall be
free of extraneous bacteria, fungi, and yeasts, as demonstrated by
microscopic examination and cultural methods.
(c) Bacterial content. (1) The number of bacteria in the concentrate
of harvested bacteria shall be estimated not later than 2 weeks after
harvest and before any treatment capable of altering the accuracy of the
estimate.
(2) The number of S. typhi bacteria in the vaccine shall not exceed
10\9\ per milliliter.
(d) Nitrogen content. The total nitrogen content of the vaccine
shall not exceed 0.035 mg./ml. for nonextracted bacteria preparations
and shall not exceed 0.023 mg./ml. for acetone-extracted bacteria
preparations.
(e) Preservative. Aqueous vaccine and the solution for
reconstitution supplied with dried vaccine shall contain a preservative.
Dried vaccine shall not contain a preservative.
[38 FR 32064, Nov. 20, 1973, as amended at 48 FR 7167, Feb. 18, 1983]
Sec. 620.12 U.S. Standard preparations.
The following U.S. Standard preparations shall be obtained from the
Center for Biologics Evaluation and Research (HFB-210), Food and Drug
Administration, 8800 Rockville Pike, Bethesda, MD 20892, for use as
prescribed in this part:
(a) Vaccine standard. The U.S. Standard Typhoid Vaccine for
determining the potency of Typhoid Vaccine.
(b) Opacity standard. The U.S. Opacity Standard for adjusting the
opacity of the suspension from which the challenge culture is prepared.
[48 FR 7167, Feb. 18, 1983, as amended at 49 FR 23834, June 8, 1984; 51
FR 15610, Apr. 25, 1986; 55 FR 11015, Mar. 26, 1990]
Sec. 620.13 Potency test.
The number of potency units per milliliter shall be estimated for
each lot of vaccine from the results of simultaneous mouse protection
tests of the vaccine under test and of the U.S. Standard Typhoid
Vaccine. At least four dilutions of each lot of vaccine shall be tested.
The test shall be performed as follows:
(a) Mice. Healthy mice shall be used, all from a single strain and
of the same sex, or an equal number of each sex in each group, with
individual weights between 13 and 16 grams. A system of randomization
shall be used to distribute the mice into the groups, with respect to
shelf position and to determine the order of challenge. A group of at
least 16 mice shall be used for each dilution of each vaccine. There
shall be at least 4 groups consisting of no less than 10 mice each for
control testing purposes, as required under paragraph (c) of this
section.
(b) Inoculation of vaccine. (1) Serial dilutions, no greater than
fivefold, of the vaccine to be tested and of the standard vaccine shall
be made in saline (0.85 percent sodium chloride solution or phosphate-
buffered saline). The mean
[[Page 75]]
effective dose (ED50) value shall be bracketed by the dilutions
used. Each mouse in each group for inoculation shall be injected
intraperitoneally with 0.5 milliliter of the appropriate dilution.
(2) The interval between inoculation of the vaccine and challenge
shall be no less than 7 days nor more than 14 days. At least 87.5
percent of the mice in each group shall survive the period between
vaccine inoculation and challenge and each mouse challenged shall appear
healthy.
(c) The challenge. (1) The challenge culture of Strain Ty 2 of S.
typhi for each test shall be taken from a batch of cultures maintained
by a method, such as freeze-drying, that retains constancy of virulence.
(2) The challenge and virulence titration doses shall be prepared as
follows: The bacteria shall be harvested from a 5- to 6-hour culture
grown at 36 deg.1 deg. C on a suitable agar medium that
shall have been seeded from a 16- to 20-hour culture grown at
36 deg.1 deg. C on a suitable agar medium, and the harvested
bacteria then shall be uniformly suspended in saline or phosphate-
buffered saline. The suspension, freed from agar particles and clumps of
bacteria and adjusted to an opacity of 10 units, shall be diluted in
saline or phosphate-buffered saline by tenfold increments. The
suspensions for the challenge and virulence titration doses shall be put
into a sterile gastric mucin preparation or other suitable virulence-
enhancing preparation. The challenge suspension shall be prepared from
whichever bacteria dilution provides about 1,000 colony forming units
for a 0.5 milliliter challenge dose. The virulence titration suspensions
shall be 101, 102, 103 dilutions, respectively, of the
challenge suspension.
(3) Each mouse inoculated with vaccine shall be injected
intraperitoneally with an 0.5 ml. dose of the challenge suspension. Each
mouse in the four groups of control mice shall be injected
intraperitoneally with an 0.5 ml. dose of the challenge suspension and
its three dilutions, respectively. The challenge dose control mice shall
be injected last. The interval between removal of the bacteria from the
culture medium and the injection of the last mouse shall not exceed 2\1/
2\ hours.
(d) Recording the results. The mice shall be observed daily for 3
days. A record shall be maintained of the number of mice that die. A
record of the number of mice that survive shall be made at the end of
the observation period.
(e) Validity of the test. The test is deemed valid if: (1) The
ED50 of the vaccine under test and the standard vaccine is between
the largest and smallest doses inoculated into the mice;
(2) The homogeneity of the dose response lines for both the vaccine
under test and the standard vaccine is acceptable;
(3) A graded protective response is obtained in relation to the
vaccine dilutions;
(4) The slopes of the dose response curves for the vaccine under
test and the standard vaccine are shown to be parallel by an appropriate
statistical method;
(5) The results of all dilutions are used to calculate the ED50
value of both the standard and test vaccine by a parallel line bioassay
method or a statistically equivalent method;
(6) The challenge dose contains approximately 1,000 colony forming
units; and
(7) The LD50 of the challenge dose contains no more than 20
colony forming units.
(f) Repeat tests. If the test does not meet the criteria prescribed
in paragraph (e) of this section, repeat tests may be performed. The
results of all tests shall be combined by geometric mean. Any test
result established as invalid under Sec. 610.1 of this chapter may be
disregarded. The determination that the vaccine meets the potency
requirements shall be made from the results of not more than four valid
tests.
(g) Estimate of the potency. The ED50 of each vaccine shall be
calculated. The protective unit value per milliliter of the vaccine
under test shall be calculated in terms of the unit value of the
standard vaccine.
(h) Potency requirements. The results of at least two separate tests
shall be included on the release protocol, required under
Sec. 620.14(c)(2), that is submitted to the Center for Biologics
[[Page 76]]
Evaluation and Research, Food and Drug Administration. The vaccine shall
have a potency of 8.0 units per milliliter. This requirement shall be
met only if the geometric mean potency for two tests is not less than
3.9 units per milliliter; or for three tests, not less than 4.4 units
per milliliter; or for four tests, not less than 4.8 units per
milliliter.
[38 FR 32064, Nov. 20, 1973, as amended at 48 FR 7167, Feb. 18, 1983; 49
FR 23834, June 8, 1984; 55 FR 11013, Mar. 26, 1990]
Sec. 620.14 General requirements.
(a) Dose. These standards are based on a human adult dose of 0.5 ml.
for a single injection and a total immunizing dose of two injections of
0.5 ml. given at appropriate intervals.
(b) Labeling. In addition to the items required by other applicable
labeling provisions of this subchapter, the package label shall state
that the vaccine contains 8 units per milliliter.
(c) Samples; protocols; official release. For each lot of vaccine,
the following material shall be submitted to the Director, Center for
Biologics Evaluation and Research (HFB-1), 8800 Rockville Pike, Bethesda
MD 20892.
(1) A sample of no less than 40 ml. of the product distributed in no
less than four containers.
(2) A protocol that consists of a summary of the history of
manufacture of each lot including all results of each test for which
test results are requested by the Director, Center for Biologics
Evaluation and Research.
(3) The product shall not be issued by the manufacturer until
written notification of official release of each filling lot of dried
vaccine and of each bulk lot of aqueous vaccine is received from the
Director, Center for Biologics Evaluation and Research.
[38 FR 32064, Nov. 20, 1973, as amended at 42 FR 27582, May 31, 1977; 48
FR 7168, Feb. 18, 1983; 48 FR 11430, Mar. 18, 1983; 49 FR 23834, June 8,
1984; 51 FR 15610, Apr. 25, 1986; 55 FR 11013 and 11015, Mar. 26, 1990]
Subpart C--Anthrax Vaccine Adsorbed
Sec. 620.20 Anthrax Vaccine Adsorbed.
The proper name of this product shall be Anthrax Vaccine Adsorbed,
which shall consist of an aqueous preparation of a fraction of Bacillus
anthracis which contains the protective antigen adsorbed on aluminum
hydroxide.
[38 FR 32064, Nov. 20, 1973, as amended at 50 FR 4137, Jan. 29, 1985]
Sec. 620.21 Production.
(a) Strain of bacteria. A nonencapsulated, nonproteolytic, avirulent
strain of Bacillus anthracis shall be used in the manufacture of anthrax
vaccine.
(b) Medium. A chemically defined medium shall be used for the
propagation of Bacillus anthracis which has protective-antigen promoting
properties that are no less effective than the protective-antigen
promoting properties of the Puziss and Wright 1095 medium as set forth
in U.S. Patent No. 3,208,909, issued September 28, 1965, which patent is
hereby incorporated by reference and deemed published herein. U.S.
Patent No. 3,208,909 has been assigned to the Federal Government and
copies will be provided to persons affected by the provisions of this
subchapter upon request to the Director, Center for Biologics Evaluation
and Research, or to the appropriate Information Center Officer listed in
45 CFR, part 5. Copies also may be obtained upon request from the U.S.
Patent Office, Washington, DC. The medium shall not contain ingredients
known to be capable of producing allergenic effects in human subjects.
(c) Propagation of bacteria. The medium shall be inoculated with a
24-hour old vegetative culture seeded from a stock suspension of spores.
The propagation culture, flushed with nitrogen, shall be incubated at
37 deg. C.plus-minus1.0 deg. C., agitated for approximately 27
hours, cooled to about 20 deg. C., the pH adjusted to
8.0plus-minus0.1 and then filtered through a sterilizing filter(s)
using nitrogen gas under pressure.
[[Page 77]]
(d) Adsorption of the protective antigen. The sterile filtrate shall
be adsorbed on sterile aluminum hydroxide gel and the recovered
precipitate shall be resuspended and diluted in sterile 0.85 percent
sodium chloride solution.
[38 FR 32064, Nov. 20, 1973, as amended at 49 FR 23834, June 8, 1984; 55
FR 11013, Mar. 26, 1990]
Sec. 620.22 U.S. Reference preparation.
The U.S. Reference Anthrax Vaccine distributed by the Center for
Biologics Evaluation and Research shall be used for determining the
potency of anthrax vaccine.
[38 FR 32064, Nov. 20, 1973, as amended at 49 FR 23834, June 8, 1984; 55
FR 11013, Mar. 26, 1990]
Sec. 620.23 Potency test.
The potency of each lot of vaccine shall be estimated from the
results of simultaneous tests of the vaccine under test and the U.S.
Reference Anthrax Vaccine. The test shall be performed as follows:
(a) Guinea pigs. Healthy guinea pigs shall be used, all from a
single strain and of the same sex, or an equal number of each sex in
each group, with individual weights between 325 and 350 grams. The diet
of the guinea pigs shall be supplemented with vitamin C throughout the
test period. At least three groups of no less than eight guinea pigs
shall be used for each vaccine and at least one group of four guinea
pigs shall be used for the challenge control.
(b) Vaccination. Serial dilutions, not greater than three-fold, of
each vaccine shall be made in 0.85 percent sodium chloride solution. The
mid-dilution of the vaccine under test shall contain that amount of
vaccine which will afford protection to approximately 50 percent of the
guinea pigs in the group vaccinated with that dilution. Each guinea pig
in the test and reference vaccine groups shall be injected
subcutaneously with 0.5 ml. of the appropriate dilution on the left side
of the abdomen and about 2 cm. from the midline. The interval between
vaccination and challenge shall be 14 days.
(c) The challenge. Each vaccinated and control guinea pig shall be
injected intracutaneously on the right side of the abdomen with 0.1 ml.
of a spore suspension of the virulent Vollum strain of Bacillus
anthracis diluted in sterile distilled water to contain 10,000 spores
per milliliter.
(d) Recording the results. The guinea pigs shall be observed daily
for 10 days and the deaths recorded. The number of survivors shall be
recorded at the end of the observation period.
(e) Validity of the test. The test shall be valid provided (1) the
protective response to each vaccine is graded in relation to the amount
of vaccine in the respective dilutions and (2) all control animals die
within 10 days.
(f) Potency requirement. The potency of the product is satisfactory
if the vaccine is no less potent than the reference. The potency of the
product is considered to be equal to the reference when (1) the average
time of death of the product-vaccinated guinea pigs is no less than the
average time of death of the reference-vaccinated guinea pigs and the
number of survivors of the product-vaccinated guinea pigs is no less
than the number of survivors of the reference-vaccinated guinea pigs, or
(2) the use of another statistical procedure, shown to be adequate for
evaluating the potency of anthrax vaccine, demonstrates that the product
is no less potent than the reference.
Sec. 620.24 General requirements.
(a) Dose. These standards are based on a single human dose of 0.5
ml. and a total primary immunizing doses of three single doses, each
given at appropriate intervals.
(b) Product characteristics. Recommendation shall be made through
appropriate labeling that the product after issue should not be frozen.
(c) Samples; protocols; official release. For each lot of vaccine,
the following material shall be submitted to the Director, Center for
Biologics Evaluation and Research, Food and Drug Administration, 8800
Rockville Pike, Bethesda, MD 20892:
(1) A protocol which consists of a summary of the manufacture of
each lot including all results of all tests for which test results are
requested by the Director, Center for Biologics Evaluation and Research.
[[Page 78]]
(2) A sample of no less than 40 milliliters of the final product
distributed in approximately equal amounts into four final containers.
(3) The product shall not be issued by the manufacturer until
written notification of official release of the lot is received from the
Director, Center for Biologics Evaluation and Research.
[38 FR 32064, Nov. 20, 1973, as amended at 42 FR 27582, May 31, 1977; 48
FR 13025, Mar. 29, 1983; 49 FR 23834, June 8, 1984; 51 FR 15610, Apr.
25, 1986; 55 FR 11013, Mar. 26, 1990]
Subpart D--Cholera Vaccine
Sec. 620.30 Cholera Vaccine.
The proper name of this product shall be Cholera Vaccine, which
shall consist of an aqueous preparation of equal parts of Ogawa and
Inaba serotypes of killed Vibrio cholerae bacteria.
[41 FR 18295, May 3, 1976]
Sec. 620.31 Production.
(a) Strains of bacteria. (1) A strain of Ogawa and a strain of Inaba
serotypes of V. cholerae shall be used in the manufacture of the
vaccine. Each serotype strain shall have been shown in controlled field
studies to yield a vaccine no less potent than vaccines prepared from
Ogawa strain 41 and Inaba strain 35A3 obtained from the Center for
Biologics Evaluation and Research.
(2) Antigenic integrity of the strains shall be verified by (i) the
agglutination of living bacteria of each serotype by cholera O Group I
antiserum; (ii) the agglutination of the Ogawa strain in monospecific
Ogawa antiserum and of the Inaba strain in monospecific Inaba antiserum;
and (iii) the absence of spontaneous agglutination of living bacteria of
either strain in 0.85 percent sodium chloride solution during incubation
for at least 5 hours at 37 deg. C.
(b) Propagation of bacteria. The culture medium for the propagation
strains shall not contain ingredients known to be capable of producing
allergenic effects in human subjects. The harvested bacteria shall be
free of extraneous bacteria, fungi, and yeasts as demonstrated by
microscopic examination and cultural methods. Bacteria of the two
serotypes shall be grown separately.
(c) Bacterial content. (1) The number of bacteria in each separate
bacterial harvest shall be determined by use of the U.S. Opacity
Standard not later than 2 hours after harvest and before treatment with
a preservative or other agent capable of altering opacity of the
bacterial suspension.
(2) The vaccine shall contain equal numbers of bacteria of the Ogawa
and Inaba serotypes, and the total number shall not exceed 8 x
109 bacteria per milliliter.
(d) Nitrogen content. The total nitrogen content of the vaccine
shall not exceed 0.3 milligram per milliliter for bacteria grown on
solid medium or 1.0 milligram per milliliter if grown in liquid medium.
In no instance shall the vaccine contain more than 0.07 milligram per
milliliter of nitrogen precipitable by the addition of an equal volume
of 10 percent trichloracetic acid.
(e) Preservative. The vaccine shall contain a preservative.
[41 FR 18295, May 3, 1976, as amended at 49 FR 23834, June 8, 1984; 55
FR 11013, Mar. 26, 1990]
Sec. 620.32 U.S. Standard preparations.
The following U.S. Standard preparations shall be obtained from the
Center for Biologics Evaluation and Research, Food and Drug
Administration, for use as prescribed in this subpart:
(a) Vaccine standard. The U.S. Standard Cholera Vaccine, Ogawa
serotype, and U.S. Standard Cholera Vaccine, Inaba serotype, shall be
reconstituted as directed for determining the potency of Cholera
Vaccine.
(b) Opacity standard. The U.S. Opacity Standard for use in
estimating the bacterial content of the vaccine and of the challenge
culture.
(c) Seed culture. Seed cultures of V. cholerae, Inaba serotype,
strain 35A3 and Ogawa serotype, strain 41, for preparation of vaccine
challenge cultures for use in the vaccine potency test.
[41 FR 18295, May 3, 1976, as amended at 49 FR 23834, June 8, 1984; 55
FR 11013, Mar. 26, 1990]
Sec. 620.33 Potency tests.
Each lot of vaccine shall be subjected to two potency tests. One
test shall determine the potency of the vaccine in comparison with the
U.S. Standard
[[Page 79]]
Cholera Vaccine, Ogawa serotype, and the other test shall determine the
potency of the vaccine in comparison with the U.S. Standard Cholera
Vaccine, Inaba serotype. At least four dilutions of each vaccine shall
be tested. Each test shall be performed as follows:
(a) Mice. Healthy mice shall be used, all from a single strain and
of the same sex, or an equal number of each sex in each group, with
individual weights between 10 and 14 grams. A group of at least 16 mice
shall be used for each dilution of each vaccine. In addition, there
shall be at least 4 groups consisting of no less than 10 mice each for
each potency test as a control for virulence titration of the challenge
suspension.
(b) Injections of vaccine. Serial dilutions, no greater than
fivefold, of the vaccine to be tested and of the appropriate serotype
standard vaccine shall be made in 0.85 percent sodium chloride solution.
The median effective dose (ED50), which is the dose of vaccine that
is expected to protect 50 percent of the animals that received the
vaccine, shall be bracketed by the dilutions used. Each mouse in each
dilution group shall receive intraperitoneally 0.5 milliliter of the
appropriate vaccine dilution. At least 87.5 percent of the mice in each
dilution group shall survive, and all surviving mice shall appear
healthy at the time of challenge.
(c) The challenge. The challenge shall be administered 12 to 16 days
after injection of the vaccine.
(1) The strains of V. cholerae for challenge shall be Ogawa 41 and
Inaba 35A3, except that V. cholerae, Inaba serotype, strain V86 may be
used instead of Inaba serotype, strain 35A3, for preparation of vaccine
challenge culture: Provided, That the source of the challenge culture
shall be identified and verified by the manufacturer as equal to that
distributed by the World Health Organization. For each test, the
challenge culture shall be taken from a batch of cultures maintained by
a method such as freeze-drying that retains constancy of virulence.
(2) The challenge and virulence titration doses shall be prepared as
follows: The bacteria for each challenge shall be harvested from a 6- to
18-hour culture grown at 36 deg.plus-minus1 deg. C, on a suitable
agar medium adjusted to pH 7.4. The harvested bacteria shall be
uniformly suspended in a diluent consisting of M/15 phosphate buffered
saline adjusted to pH 7.4 and shall contain 0.1 to 0.2 percent gelatin.
The suspension shall be free from agar particles and clumps of bacteria.
The suspension shall be adjusted to an opacity of 10 units, and diluted
in tenfold increments using the same diluent. The suspensions for the
challenge and virulence titrations shall be suspended in a 5 to 10
percent sterile gastric mucin preparation adjusted to pH 7.4. The
challenge suspension shall be prepared from whichever bacterial dilution
provides the required median lethal dose (LD50) for a 0.5
milliliter challenge dose. The LD50 is the dose of the challenge
suspension that is expected to kill 50 percent of the animals that
received the challenge. The virulence titration suspensions shall
consist of the challenge suspension and at least three dilutions of the
challenge suspension calculated to bracket the LD50 value.
(3) At least 16 surviving mice, randomly selected from each dilution
group that received vaccine, shall be inoculated intraperitoneally with
a 0.5-milliliter dose of the challenge suspension. Mice in each of the
four groups of control mice used for the virulence titration of the
challenge suspension shall be inoculated intraperitoneally with a 0.5-
milliliter dose of the challenge suspension and its respective
dilutions. The challenge dose control mice shall be inoculated last. The
interval between removal of the bacteria from the culture medium and the
inoculation of the last mouse shall not exceed 2\1/2\ hours.
(d) Recording the results. The mice shall be observed daily for 2
days following challenge. A daily record shall be maintained of the
number of mice that die. A record of the number of mice that survive
shall be made at the end of the observation period.
(e) Validity of the test. The test is valid provided: (1) The
ED50 value of the vaccine under test and the standard vaccine is
between the largest and smallest doses inoculated into the mice;
[[Page 80]]
(2) The homogeneity of the dose response lines for both the vaccine
under test and the standard vaccine is acceptable;
(3) The log-dose response lines for the vaccine under test and the
standard vaccine are shown to be parallel by an appropriate statistical
method;
(4) The results of all dilutions shall be used to calculate the
ED50 value of both the standard and test vaccine by a parallel line
bioassay method or a method statistically equivalent;
(5) The challenge dose contains between 100 and 10,000 LD50
doses; and
(6) The LD50 value of the challenge suspension contains no more
than 10,000 colony-forming units determined by plate count.
(f) Repeat tests. Repeat tests need be performed only on the
serotype which failed to meet the potency requirements prescribed in
paragraph (h) of this section. The results of each test on each serotype
meeting the criteria in paragraph (e) of this section shall be combined
by means of a geometric mean. The determination that the vaccine meets
the potency requirements shall be made from the results of not more than
three valid tests on each serotype.
(g) Estimate of the potency. The ED50 value of each vaccine
shall be calculated. The protective unit value of each serotype per
milliliter of the vaccine under test shall be calculated in terms of the
unit value of the corresponding standard vaccine.
(h) Potency requirements. The vaccine shall have a potency of not
less than 8 units per serotype per milliliter. This requirement shall be
met only if the potency for a single test is not less than 4.4 units per
serotype per milliliter, or for two tests not less than 5.3 units, or
for three tests not less than 5.7 units.
[41 FR 18295, May 3, 1976, as amended at 41 FR 46587, Oct. 22, 1976]
Sec. 620.34 Mouse toxicity test.
The final vaccine shall be demonstrated to be free from toxicity by
the following test: A group of no less than 10 and no more than 40 mice,
each mouse weighing 14 to 16 grams, shall have free access to food and
water at least 2 hours before injection and throughout the test period.
The group weight of the mice shall be determined immediately before
injection. Each mouse shall be injected intraperitoneally with a test
dose of 0.5 milliliter of undiluted vaccine. The group weight of the
mice shall be determined again at the end of 72 hours. The 72-hour
average weight per mouse shall be no less than the average weight per
mouse immediately preceding the injection. No more than 5 percent of the
total number of mice used may die during the test period; however,
neither death nor significant toxic signs attributable to the vaccine
shall result.
[41 FR 18295, May 3, 1976]
Sec. 620.35 General requirements.
(a) Freezing prohibition. Cholera Vaccine shall not be frozen at any
time.
(b) Dose. These standards are based on a total immunizing dose of
two injections of 0.5 milliliter and 1.0 milliliter, respectively, given
at intervals specified in the manufacturer's labeling.
(c) Date of manufacture. The date of manufacture shall be the date
of initiation of the last valid potency test for the Ogawa serotype or
the Inaba serotype, whichever date is earlier.
(d) Labeling. In addition to the applicable labeling provisions of
this chapter, the package label shall bear the following: (1) A
statement that the vaccine contains 8 units of each serotype antigen per
milliliter.
(2) The statement, ``DO NOT FREEZE''.
(3) The statement, ``SHAKE WELL''.
(e) Samples; protocols; official release. For each lot of vaccine,
the following material shall be submitted to the Director, Center for
Biologics Evaluation and Research, Food and Drug Administration, 8800
Rockville Pike, Bethesda, MD 20892.
(1) A sample consisting of no less than 40 milliliters of the
product. The sample may be in the final container or from the vaccine
bulk lot.
(2) A protocol which consists of a summary of the history of
manufacture of each lot including all results of each test for which
test results are requested by the Director, Center for Biologics
Evaluation and Research,
[[Page 81]]
Food and Drug Administration. The raw data and results from each potency
test performed shall be included.
(3) The product shall not be issued by the manufacturer until
written notification of official release of the lot is received from the
Director, Center for Biologics Evaluation and Research.
[41 FR 18295, May 3, 1976, as amended at 42 FR 27582, May 31, 1977; 49
FR 23834, June 8, 1984; 51 FR 15610, Apr. 25, 1986; 55 FR 11013, Mar.
26, 1990]
Subpart E--Bacillus of Calmette and Guerin (BCG) Vaccine
Source: 44 FR 14545, Mar. 13, 1979, unless otherwise noted.
Sec. 620.40 BCG Vaccine.
(a) Proper name and definition. The proper name of this product is
BCG Vaccine. The product is defined as a freeze-dried preparation
containing viable bacteria of the Bacillus of Calmette and Guerin, which
is an attenuated strain of Mycobacterium bovis.
(b) Criteria for an acceptable strain. The source of the BCG strain
used in the manufacture of any lot of the final product must be
identified by complete historical records.
(1) Seed lot system. The BCG strain must be maintained in the form
of a primary seed lot that is to be the basic material from which all
secondary seed lots are prepared. Production of BCG Vaccine may be from
either primary or secondary seed lots. Each seed lot must be stored in
either a freeze-dried state at -20 deg. C or colder, or in a frozen
state at -70 deg. C or colder.
(2) Freedom from virulence. The BCG strain is demonstrated to be
incapable of producing progressive tuberculosis in guinea pigs tested as
prescribed in Sec. 620.45, except that no fewer than 48 guinea pigs must
be used to test the primary seed lot and no fewer than 12 guinea pigs
must be used to test each secondary seed lot. At least two-thirds of the
animals must survive the observation period of no less than 6 months.
(3) Induction of tuberculin sensitivity in guinea pigs. Each of at
least 10 guinea pigs is to be injected with 1 human dose of BCG Vaccine
and, within 4 to 6 weeks after vaccination, skin tested with tuberculin.
At least 80 percent of the guinea pigs tested must develop tuberculin
sensitivity, as prescribed in Sec. 620.44(b)(3)(ii).
(4) Clinical information. Clinical data must establish that the BCG
strain is safe and induces tuberculin sensitivity. After having passed
all laboratory tests prescribed for BCG Vaccine, each primary and
secondary seed lot of vaccine must be tested for its ability to induce
sensitivity in tuberculin-negative persons. Only those persons tested by
injection of 5 U.S. Tuberculin Units, Purified Protein Derivative, by
the Mantoux technique and found negative in this test are to be selected
for clinical trials. At least 100 tuberculin-negative persons must be
included in the test of the primary seed lot, and at least 20
tuberculin-negative persons must be included in the test of each
secondary seed lot. Within 6 to 8 weeks after BCG vaccination, the
vaccinees must be tested for tuberculin reactivity by injecting not more
than 10 U.S. Tuberculin Units, Purified Protein Derivative, by the
Mantoux technique. The test is considered satisfactory if a least 90
percent of those persons from each group develop tuberculin reactivity
as indicated by an induration reaction of at least 5 millimeters in
diameter.
Sec. 620.41 Establishment and personnel requirements.
In addition to the applicable requirements of Secs. 600.10 and
600.11 of this chapter, the following practices and procedures are
required:
(a) Isolation of BCG unit. (1) A BCG unit is defined as the space
used for storage of primary and secondary seed cultures and for vaccine
preparation, including culture maintenance, media inoculation for
propagation, harvesting, filling into final containers, sealing of final
containers, media production, and cleaning and sterilization of
glassware. For purposes of these additional standards, the space used
for incubation of bulk and final container sterility tests, tests to
determine the numbers of colony-forming units, animal tests, and
necropsies are not part of the BCG unit.
(2) The BCG unit must be completely isolated from other production
and surrounding areas and must be situated
[[Page 82]]
and designed to prevent contamination of the product. It must have a
separate facility for ventilation, designed to prevent contamination of
the product. The facilities for water supply and sewage and trash
disposal must be designed to prevent microbial contamination of the BCG
unit. The equipment used in BCG Vaccine production must remain in the
BCG unit at all times.
(3) Microbial controlled areas must be available for handling the
BCG cultures. No cultures of microorganisms other than the BCG
production strain are permitted in the BCG unit. No animals are
permitted in the BCG unit. All tests necessary for the control of the
vaccine, in which contaminating microorganisms may be cultured, or in
which animals are used, must be conducted in space physically separated
from the BCG unit.
(b) Restrictions on personnel. (1) A staff specially trained in
maintaining the seed cultures, propagating the cultures, preparing the
vaccine, and filling the vaccine into final containers shall be employed
in the production of the BCG Vaccine. Such personnel shall not work with
other infectious agents in any laboratory at any time and shall not be
exposed to a known risk of tuberculosis. Within 30 days before
employment in the BCG unit, each person working in the unit shall have
had a medical examination, including a tuberculin skin test with 5 U.S.
Tuberculin Units, Purified Protein Derivative, by the Mantoux procedure,
and a chest X-ray. No person who has had a history of tuberculosis or
mycobacterial disease is permitted in the BCG unit. There must be
periodic medical examinations of BCG unit personnel, including X-ray
examinations, of sufficient frequency to detect the appearance of early
active tuberculosis. Repeated tuberculin skin testing of staff who are
negative to tuberculin may be used as an additional diagnostic aid in
isolating any potential source of tuberculosis exposure. If a person
working in the BCG unit develops active tuberculosis, (i) the entire
staff shall be examined for possible tuberculosis infection, (ii) all
current vaccine preparations and all cultures with which the person may
have come into contact since his or her last satisfactory medical
examination, except cultures sealed before that examination, must be
discarded, and (iii) the BCG unit and all equipment with which the
person may have come in contact must be decontaminated.
(2) Personnel shall wear protective clothing and use protective
devices to the extent necessary to protect the product from
contamination.
(3) Any person not assigned to the BCG unit shall not be allowed
into the BCG unit at any time unless a medical examination shows the
person to be free from mycobacterial disease.
Sec. 620.42 Production.
(a) BCG inoculum. The inoculum of BCG used for seed lot or
production of final lot in seed buildup must have been removed from the
preceding seed lot in accordance with the following passage and time
schedule:
(1) No more than 3 passages from primary to secondary seed lot
within a 2-month period.
(2) If no secondary seed lot is used, no more than 9 passages from
primary seed lot to final lot within a 6-month period.
(3) No more than 9 passages from secondary seed lot to final lot
within a 6-month period.
(b) Propagation of bacteria. The culture medium for propagation of
BCG Vaccine must not contain ingredients known to be capable of
producing allergenic effects in humans or of causing the bacteria to
become virulent for guinea pigs. The growth in each container must be
examined visually, and only those cultures that have the typical growth
pattern characteristic of BCG are to be used in a vaccine.
(c) Colony-forming units (CFU) before and after freeze-drying. Each
lot of BCG Vaccine must be tested to determine the number of CFU per
individual final container both before and after freeze-drying, by the
method prescribed in Sec. 620.44(a). The upper and lower limits of the
viable count are to be established by the manufacturer of the vaccine
for the particular route of administration recommended and must be
specified in the license application. The loss in viability after drying
must not exceed 90 percent.
[[Page 83]]
Sec. 620.43 Reference BCG Vaccine.
A reference BCG Vaccine, for use in determining the validity of the
test for colony-forming units, is to be obtained from the Director,
Center for Biologics Evaluation and Research, Food and Drug
Administration, 8800 Rockville Pike, Bethesda, MD 20892.
[44 FR 14545, Mar. 13, 1979, as amended at 49 FR 23834, June 8, 1984; 51
FR 15610, Apr. 25, 1986; 55 FR 11013, Mar. 26, 1990]
Sec. 620.44 Potency tests.
(a) Colony-forming units (CFU). The number of CFU must be determined
on the contents of each of at least 10 individual final containers of
each lot of BCG Vaccine. Of the 10 or more individual final containers,
the contents of at least 5 before, and an equal number after, freeze-
drying must be tested. Final containers of the freeze-dried vaccine are
to be reconstituted as for human use with the diluent recommended by the
manufacturer. The number of CFU to be reported for each lot of BCG
Vaccine must be determined only from test tubes containing between 10
and 50 CFU. Dilutions must be made as follows:
(1) Dilutions are made from an appropriate volume of the liquid
vaccine before freeze-drying or the reconstituted vaccine after freeze-
drying. Appropriate dilutions are made with modified Youman's medium
specified in paragraph (a)(4) of this section, up to a point where
subsequent serial half-log dilutions will result in at least 1 tube
containing between 10 and 50 CFU.
(2) Serial half-log dilutions are made in 16 x 125 millimeter screw-
capped test tubes into which 4.5 milliliter aliquots of the diluent
prescribed in paragraph (a)(4) of this section have been dispensed. Two
milliliters of thoroughly mixed vaccine are added to the first tube of
the half-log series, mixed thoroughly, and 2.0 milliliters from this
tube are transferred to the next tube in the series. The process of
mixing and serially transferring 2.0 milliliters is repeated through
each consecutive tube and 2.0 milliliters are discarded from the last
tube.
(3) After the serial half-log dilutions are completed, 0.5
milliliter of 1.5 percent agar solution that has been cooled to 42 deg.
C is quickly added, where necessary, to make a final concentration of
0.15 percent agar, and the contents of the tubes are thoroughly mixed.
After mixing, all tubes are incubated at 35 deg. to 37 deg. C for 3 to 4
weeks.
(4) The composition of modified Youman's medium with bovine albumin
is as follows:
Asparagine................................ 5.0 grams.
Monopotassium phosphate (KH2PO4).......... Do.
Potassium sulfate (K2SO4)................. 0.5 grams.
Magnesium citrate......................... 1.5 grams.
Monosodium glutamate...................... 19.0 grams.
Glycerine................................. 20.0 milliliters.
Distillled water q.s. to.................. 900.0 milliliters.
One hundred milliliters of 5-percent aqueous solution of bovine
albumin that has been sterilized by filtration are added to the Youman's
medium to produce a final concentration of 0.5 percent of bovine
albumin. The pH is adjusted to 7.0 with 5N sodium hydroxide.
(b) Intradermal guinea pig test. Two or more guinea pigs, each
weighing no less than 250 grams, must be injected intradermally in 4
different sites with the following amounts and dilutions of each lot of
BCG Vaccine:
(1) Vaccine intended for intradermal injection is reconstituted as
for human use with the diluent recommended by the manufacturer. One-
tenth milliliter of reconstituted vaccine and 0.1 milliliter each of
three ten-fold dilutions (1:10, 1:100, and 1:1000) of the reconstituted
vaccine are injected into the guinea pigs. The diluent for the ten-fold
dilutions is isotonic solution for injection.
(2) Vaccine intended for percutaneous injection into humans is
reconstituted with the diluent recommended by the manufacturer so that
at least one human dose (estimated to be within a range of from 1 to
33 x 105 CFU) is contained in 0.1 milliliter. A narrower range of
CFU is determined for each specific vaccine by the manufacturer and
specified in the license application. One-tenth milliliter of the
selected dose of vaccine and 0.1 milliliter each of three ten-fold
dilutions (1:10, 1:100, and 1:1000) are injected into the guinea pigs.
The diluent for the ten-fold dilutions is an isotonic solution for
injection.
(3) The lot of vaccine is satisfactory if:
[[Page 84]]
(i) At the end of 2 to 4 weeks after BCG vaccination, nodules have
developed that are graded in relation to the amount of the test dose,
with the largest dose inducing a nodule of from 4 to 10 millimeters in
diameter and the smallest dose inducing essentially no nodule, and all
observations are read on the same day;
(ii) By the end of 4 to 6 weeks after BCG vaccination, each guinea
pig shows a degree of sensitivity such that an intradermal injection of
no greater than 25 U.S. Tuberculin Units, Purified Protein Derivative,
in 0.1 milliliter will induce an erythematous reaction at least 10
millimeters in diameter within 18 to 24 hours; and
(iii) At the end of the test period, each guinea pig is weighed and
each shows a weight increase.
(c) Induction of tuberculin sensitivity in tuberculin-negative
humans. At least once annually, no less than one lot of BCG Vaccine that
has satisfied all requirements and has been released by the Center for
Biologics Evaluation and Research must be tested for its ability to
induce sensitivity in 20 persons negative to Tuberculin, Purified
Protein Derivative, as prescribed in Sec. 620.40(b)(4). The results of
these tests must be sent to the Director, Center for Biologics
Evaluation and Research, as they are completed.
[44 FR 14545, Mar. 13, 1979, as amended at 49 FR 23834, June 8, 1984; 55
FR 11013, Mar. 26, 1990]
Sec. 620.45 Test for freedom from virulent mycobacteria.
(a) Each lot of BCG Vaccine must be tested to determine that it does
not contain virulent mycobacteria. The test must be performed using at
least 6 guinea pigs, each weighing between 250 and 300 grams. Vaccine
intended for intradermal injection in humans must be tested by injecting
into guinea pigs the number of bacteria contained in at least 50 human
doses. Vaccine intended for percutaneous use in humans must be tested by
injecting into guinea pigs 50 times the number of bacteria estimated to
be introduced parenterally into humans by the recommended procedure. The
vaccine for all tests must be inoculated subcutaneously or
intramuscularly into the guinea pigs. All animals that die during the
observation period must be examined postmortem. All animals that survive
the observation period must be sacrificed and examined post mortem. The
lot passes the test if at least two-thirds of the animals on test
survive an observation period of not less than 6 weeks, and if the post-
mortem examination reveals no evidence of tuberculosis in any of the
test animals.
(b) If any virulent mycobacteria are found in any lot of BCG
Vaccine, whether or not the manufacturer intends to submit samples and
protocols of this lot to the Center for Biologics Evaluation and
Research for release, the following actions must be taken:
(1) In addition to the requirements of Secs. 600.12 and 600.14 of
this chapter, the manufacturer shall immediately report by telephone,
telegraph, or cable the finding of virulent mycobacteria to the
Director, Center for Biologics Evaluation and Research.
(2) All production and distribution of lots of BCG Vaccine produced
from the same secondary seed lot as the contaminated lot of BCG Vaccine
must be discontinued. If no secondary seed lot is used the same
requirements apply to the primary seed lot.
(3) The manufacturer shall conduct a thorough and prompt
investigation concerning the failure of the lot to meet the required
safety and purity specifications, including retesting the suspect lot
and the source secondary seed lot (or primary seed lot, if no secondary
seed lot is used) and shall undertake a thorough review of all
manufacturing records and procedures to determine the probable cause of
the failure.
(4) A written record of the investigation, including the retest
results, must be submitted to the Director, Center for Biologics
Evaluation and Research.
(5) Neither production nor distribution of BCG Vaccine may be
resumed until the manufacturer is notified in writing by the Director,
Center for Biologics Evaluation and Research, that such activity may be
resumed.
[44 FR 14545, Mar. 13, 1979, as amended at 49 FR 23834, June 8, 1984; 55
FR 11013, Mar. 26, 1990]
[[Page 85]]
Sec. 620.46 General requirements.
(a) Dose. These standards are based on (1) vaccine intended for
intradermal injection in a single human immunizing dose of 0.1
milliliter and (2) vaccine intended for percutaneous injection in a
single skin application through which inoculation is made by a multiple
puncture device.
(b) Date of manufacture. The date of manufacture is the date of
initiation of the last valid determination for CFU after freeze-drying.
Sec. 620.47 Labeling.
In addition to conforming to the applicable requirements of
Secs. 610.60, 610.61, and 610.62 of this chapter, the package label must
bear the following information:
(a) Specification of the route of administration.
(b) A statement that the vaccine contains live bacteria and should
be protected against exposure to light.
(c) A statement that the vaccine must be administered within 8 hours
after reconstitution, and that reconstituted vaccine not used within 8
hours must be discarded.
Sec. 620.48 Samples; protocols; official release.
(a) For each lot of vaccine, the following materials must be
submitted to the Director, Center for Biologics Evaluation and Research,
Food and Drug Administration, 8800 Rockville Pike, Bethesda, MD 20892.
(1) Samples and diluent that will provide at least 20 milliliters
when the samples are reconstituted as recommended in the package insert
by the manufacturer of the vaccine.
(2) A protocol that consists of a complete summary of the
manufacture of each lot, including all results of each test required by
all applicable regulations. If the protocol is not included in the
shipment of the samples, it must be sent promptly to the Director,
Center for Biologics Evaluation and Research, Food and Drug
Administration, 8800 Rockville Pike, Bethesda, MD 20892.
(b) The BCG Vaccine must not be issued by the manufacturer until
written notification of official release is received from the Director,
Center for Biologics Evaluation and Research, Food and Drug
Administration.
[44 FR 14545, Mar. 13, 1979, as amended at 49 FR 23834, June 8, 1984; 51
FR 15610, Apr. 25, 1986; 55 FR 11013, Mar. 26, 1990]