[Federal Register Volume 60, Number 89 (Tuesday, May 9, 1995)]
[Rules and Regulations]
[Pages 24547-24549]
From the Federal Register Online via the Government Publishing Office [www.gpo.gov]
[FR Doc No: 95-11294]
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DEPARTMENT OF AGRICULTURE
9 CFR Part 113
[Docket No. 93-071-2]
Viruses, Serums, Toxins, and Analogous Products; Detection of
Extraneous Agents by the Fluorescent Antibody Technique
AGENCY: Animal and Plant Health Inspection Service, USDA.
ACTION: Final rule.
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SUMMARY: We are amending 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 amendment allows the use of alternative fluorochromes
that may be conjugated to an antibody, revises the list of extraneous
agents to be tested for, and includes extraneous agents for which
equine cells are to be tested. In addition, the word ``agent'' is
replaced with the word ``virus'' since this is the agent being tested
for. The amendment is necessary to update the requirements related to
the testing for extraneous viruses.
EFFECTIVE DATE: June 8, 1995.
FOR FURTHER INFORMATION CONTACT:
Dr. David A. Espeseth, Deputy Director, Veterinary Biologics, BBEP,
APHIS, 4700 River Road Unit 148, Riverdale, MD 20737-1237, (301) 734-
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 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
[[Page 24548]] 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 become necessary.
On March 21, 1994, we published in the Federal Register (59 FR
13257-13259, Docket No. 93-071-1) a proposal to amend the regulations
by revising Sec. 113.47 to allow the use of additional fluorescent
stains in the testing of extraneous viruses by the fluorescent antibody
technique. 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. The
amendment thus allows for the use of alternative fluorochromes in such
tests. The term ``fluorescein-conjugated antibody'' is replaced with
``fluorochrome-conjugated antibody'' everywhere it appears in
Secs. 113.47 and 113.52(b)(2) (i) and (ii).
The current regulations in Sec. 113.47 lists the specific
extraneous viruses against which antibodies are used in the testing of
certain types of cells. We have 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 specific viruses before such use is approved.
We solicited comments concerning our proposal for a 60-day comment
period ending April 20, 1994. We received one comment by that date from
a manufacturer of veterinary biological products. We carefully
considered the comment which is discussed below.
The comment was in two parts The first was that the fluorescent
antibody (FA) test was not as sensitive for detecting bluetongue virus
as seroconversion in sheep, and the use of the FA test would require
firms to introduce this organism into their testing facility. The
second was that testing of bovine cells for bovine respiratory
syncytial virus was also a new requirement.
In response to the comment, the Animal and Plant Health Inspection
Service has found that the FA test is a sensitive, specific, and
accurate test for the detection of extraneous viruses in seeds, cells
and ingredients of animal origin. When used with the required controls,
the FA test detects the presence of specific antigens indicative of the
presence of specific extraneous viruses. The use of positive and
negative controls with the FA test eliminates the incidence of false
positives and false negatives. The FA test also has the advantage of
relieving the firms from the need to locate and maintain seronegative
sheep.
APHIS acknowledges that requiring firms to test for the bluetongue
virus will result in the introduction of the agent onto their premises.
This should, however, pose no greater problem to the firms than
introducing other viruses for the purpose of conducting these tests. If
no firm's biosecurity is adequate to contain such viruses as those
causing bovine virus diarrhea, canine parvoviral diarrhea, or
pseudorabies, it should be able to contain bluetongue virus. Further,
there is no threat of human disease from the use of bluetongue virus in
association with this test as bluetongue virus is not known to affect
humans.
With respect to the requirement for testing bovine cells for bovine
respiratory syncytical virus, APHIS has determined that this virus may
be present as an extraneous agent in cells of bovine origin. One of the
major purposes of the rule is to update the list of extraneous viruses
for which seeds, cells, and ingredients of animal origin are tested.
The new requirements are based on current knowledge of animal diseases.
No changes are made to the regulations in response to the commenter.
Therefore, based on the rationale set forth in the proposed rule
and in this document, we are adopting the provisions of the proposal as
a final rule with one conforming change in Sec. 113.300(c)(1).
Executive Order 12866 and Regulatory Flexibility Act
This final rule has been reviewed under Executive Order 12866. This
rule has been determined to be not significant for the purposes of
Executive Order 12866, and, therefore, has not been reviewed by the
Office of Management and Budget.
These amendments should not have a significant economic impact on
manufacturers since they will broaden the range of fluorochrome stains
that may be used in conducting the fluorescent antibody test and will
revise the list of extraneous agents for which various cell types are
to be tested with the fluorescent antibody technique. The amendments
will thus remove outdated requirements and provide flexibility in the
types of antibody that may be used in tests for extraneous agents. On
balance, therefore, there should be no net increase in testing
requirements over the current requirements.
Under these circumstances, the Administrator of the Animal and
Plant Health Inspection Service has determined that this action will
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 final rule has been reviewed under Executive Order 12778,
Civil Justice Reform. This rule: (1) Preempts all State and local laws
and regulations that are inconsistent with this rule; (2) has no
retroactive effect; and (3) does not require administrative proceedings
before parties may file suit in court challenging this rule.
Paperwork Reduction Act
This rule contains no information collection or recordkeeping
requirements under the Paperwork Reduction Act of 1980 (44 U.S.C. 3501
et seq.).
List of Subjects in 9 CFR Part 113
Animal biologics, Exports, Imports, and Reporting and recordkeeping
requirements.
Accordingly, 9 CFR part 113 is amended as follows:
PART 113--STANDARD REQUIREMENTS
1. The authority citation for part 113 continues 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 is 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 [[Page 24549]] last subculturing, shall be processed and stained
with the appropriate antiviral fluorochrome-conjugated antibody as
specified in paragraph (b) of this section.
(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
cm\2\.
(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 virsus;
(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 cells 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), is 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
fluorchrome-conjugated antibody specific to the species of origin of
the MSC.
* * * * *
Secs. 113.51, 113.52 and 113.53 [Amended]
4. In the following places, the word ``agents'' are 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).
5. In Sec. 113.300, paragraph (c)(1), the term ``fluorescein'' is
removed and the term ``fluorochrome'' added in its place.
Done in Washington, DC, this 1st day of May 1995.
B. Glen Lee,
Acting Administrator, Animal and Plant Health Inspection Service.
[FR Doc. 95-11294 Filed 5-8-95; 8:45 am]
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