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Guinea Pigs
Single inoculation regime
Repeat inoculation regime
Rabbits
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PROCEDURE FOR PRODUCING ANTI-FMDV SERA
Guinea Pigs
1. Single inoculation regime
Guinea pigs, 600-700 g in weight, are inoculated subcutaneously with an
emulsion of 10 µg of inactivated FMDV antigen in sterile Tris buffer,
pH 7.6., (0.05 M Tris, 0.1 M NaCl, 0.1% sodium azide), emulsified in an
equal volume of Freund's complete adjuvant (FCA), 1 ml per guinea pig
[see Appendix 1 for production of purified virus antigen].
Ensure emulsion is thoroughly mixed.
This can be done by repeatly passing through a small bore needle until
an emulsion is formed that does not disperse when a drop is placed in
water.
The animals are killed and exsanguinated after 28 days.
2. Repeat inoculation regime
Guinea pigs are inoculated subcutaneously with an emulsion of 5 µg of
inactivated FMDV antigen in Tris buffer mixed with an equal volume of
FCA, 1 ml per guinea pig. The animals are boosted at 28 days as above,
this time using Freund's incomplete adjuvant (FIA).
The animals are killed and exsanguinated 10-12 days later.
Guinea pig sera can be heat-inactivated at 56°C for 30 min, pooled as
required, Seitz filtered and stored at -20°C.
Rabbits
Rabbits are inoculated subcutaneously and intra-muscularly with an emulsion
of 15 µg of inactivated FMDV antigen, plus FCA, at each of several sites,
1 ml per rabbit. Each rabbit is boosted 28 days later, as for the repeat
inoculation regime for guinea pigs, using FIA.
The animals are killed and exsanguinated 10-12 days later.
Rabbit antiserum for the antigen ELISA is usually not heat-inactivated.
It is often worthwhile to take test bleeds from animals near to 28 (and
40 days for the repeat inoculation regime) to obtain an approximate evaluation
of antibody response. If indications are that the antiserum is poor it
may be advisable to use another antigen preparation for re-inoculation,
and/or postpone killing and exsanguination of the animals.
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