[Code of Federal Regulations]
[Title 9, Volume 1]
[Revised as of January 1, 2002]
From the U.S. Government Printing Office via GPO Access
TITLE 9--ANIMALS AND ANIMAL PRODUCTS
CHAPTER I--ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF
PART 113--STANDARD REQUIREMENTS--Table of Contents
Sec. 113.53 Requirements for ingredients of animal origin used for production of biologi cs.
Each lot of ingredient of animal origin which is not subjected to
heat sterilization or other sterilization methods acceptable to Animal
and Plant Health Inspection Service (APHIS), such as, but not limited to
serum and albumin, used to prepare a biological product shall be tested
as prescribed in this section by the licensee or a laboratory acceptable
to VS. Results of all tests shall be recorded by the testing laboratory
and made a part of the licensee's
records. A lot of ingredient found unsatisfactory by any prescribed test
shall not be used to prepare a biological product. A serial of
biological product shall not be released if produced using an ingredient
that is found unsatisfactory by any prescribed test.
(a) Samples of each lot of ingredient of animal origin which is not
subjected to heat sterilization, used to prepare a biological product
shall be shown free of mycoplasma by the method prescribed in
(b) Samples of each lot of ingredient or animal origin which is not
subjected to heat sterilization of other sterilization methods
acceptable to APHIS used to prepare a biological product shall be shown
free of bacteria and fungi as prescribed in Sec. 113.26.
(c) Samples of each lot of ingredient of animal origin, except
porcine trypsin, which is not subjected to heat sterilization or other
viricidal procedure acceptable to APHIS used in the preparation of
biological products shall be tested as prescribed in this paragraph;
(1) Monolayers at least 75 cm\2\ of Vero (African green monkey
kidney) cell line and of primary cells or a cell line of the same
species of origin as the ingredient shall be used in the test. Cell
lines used shall have been found satisfactory when tested as prescribed
in Sec. 113.52 and primary cells used shall have been found satisfactory
when tested as prescribed in Sec. 113.51.
(2) At least 3.75 ml or 15 percent of the ingredient shall be used
in the growth medium for the preparation of at least 75 cm\2\ test
monolayers. The ingredient shall also be used in the growth medium when
monolayers are subcultured. If the ingredient being tested is cytotoxic
when tested in this manner, other procedures may be used if approved by
(3) The test monolayers shall be maintained for at least 21 days.
(4) Cells shall be subcultured at least two times during the
maintenance period. All but the last subculture shall result in at least
one new monolayer of at least 75 cm\2\. The last subculture shall meet
the minimum area requirements specified in Secs. 113.46 and 113.47.
(5) Monolayers shall be examined regularly throughout the 21-day
maintenance period for evidence of cytopathogenic agents. If evidence of
a cytopathogenic agent is found, the ingredient is unsatisfactory.
(6) At the conclusion of the 21-day maintenance period, monolayers
shall be tested for:
(i) Cytopathogenic and/or hemadsorbing agents as prescribed in
Sec. 113.46; and
(ii) Extraneous viruses by the fluorescent antibody technique as
prescribed in Sec. 113.47.
(d) Each lot of porcine trypsin which has not been treated to
inactivate porcine parvovirus (PPV) in a manner acceptable to VS shall
be tested for PPV as prescribed in this paragraph.
(1) Not less than 5.0 grams of trypsin shall be dissolved in a
volume of suitable diluent sufficient to fill a centrifuge angle head.
After centrifuging for 1 hour at 80,000 x g, the pellet material shall
be reconstituted in distilled water and inoculated into a flask
containing 75 cm\2\ of a 30 to 50 percent confluent monolayer culture of
primary porcine cells or a porcine cell line of proven equal PPV
susceptibility. An additional flask of cells shall be held as a negative
(2) The test and control monolayers shall be maintained for at least
14 days and subcultured at least once during the maintenance period.
(3) At the end of the 14-day maintenance period, and 4 to 7 days
after the last subculturing, monolayers shall be tested for the presence
of porcine parvovirus by the fluorescent antibody technique as
prescribed in Sec. 113.47(c).
(e) A sample of serum from each donor horse used to produce a lot of
equine serum used in the preparation of biological products recommended
for use in horses shall be tested at a laboratory approved by Animal and
Plant Health Inspection Service using the Coggins test for equine
infectious anemia antibodies. If antibodies to equine infectious anemia
are found, the lot of serum is unsatisfactory.
[50 FR 442, Jan. 4, 1985; 50 FR 3316, Jan. 24, 1985, as amended at 56 FR
66784, Dec. 26, 1991; 60 FR 24549, May 9, 1995]