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Standard 70 days Immunization Serum Production Protocol - Rabbit

Standard 70 days Immunization Serum Production Protocol - Rabbit

This is our standard 70 serum production protocol, If the antigen is provided by the customer: 2.0-2.5 mg of peptide or 2.0-2.5 mg of protein is required to immunize two rabbit. .  The concentration should be >0.4 mg/ml, and the antigen solution must not contain organic solvents.

Note: Client-design production protocol also available at additional charge

  • Species – New Zealand White Rabbits

  • Average age of rabbits - 6 months

  • Average Weight of Rabbits – 5-6 lbs

Immunization/Boost

The antigen/adjuvant mixture is inoculated multiple site subcutaneously along the back and intramuscularly in the hind limbs. We use no more than 0.1ml of the mixture per site. If we have 1.0 ml of antigen, we mix it with 1.0ml of Complete Freund’s Adjuvant for initial inoculation and 1.0ml of Incomplete Freund’s Adjuvent for the following injections. This means we have 2.0ml of mixture, and therefore, we will do 10 subcutaneous sites and 10 intramuscular sites.

Day  Procedure
0 PreBleed/ Initial Inoculation(200mg of immunogen at 0.5ml of conjugate solution mixed with an equal volume of Complete Freund’s Adjuvent)  100 ml is injected into each site  - 10 sites total.
14 Boost (200mg of immunogen at 0.5ml of conjugate solution mixed with an equal volume of Incomplete Freund’s Adjuvent)  100ml is injected into each site  - 10 sites total.
28 Boost (As on day 14 above)
42 Boost (As on day 14 above)/ Production bleed (Approximately 20 ml Sera)
56 Boost (As on day 14 above)/ Production bleed (Approximately 20 ml Sera)
70  Production Bleed (Approximately 20 ml Sera)

The frequency of the production bleeds is based upon the size, health, hematocrit, etc. of the host species and the requirements of the investigator.

Care of Animals:

Each animal is housed in an individual cage. There are as many as 1,000 cages in a building so the animals are not isolated from each other. The rabbits are fed a premeasured, pelleted ration once each day and have constant access to water (from a well on the property). The buildings are heated in the winter and cooled in the summer. The temperature is kept between 68oF and 85oF. The lights in the housing facilities are on a timer so that the rabbits have 12 hours of light and darkness each day. There is a regular schedule of cleaning the cages, feeders, dropping trays, water lines and lixits, floors, walls, ceilings, etc. We follow NIH and USDA guidelines for all care and maintenance of the animals. We have consulting veterinarians who are available 24 hours a day, 365 days a year for any care or health issues, which may arise with the animals.

Handling of Blood from Rabbits:

Once the rabbit is bled, the whole blood is allowed to set at room temperature for about 2 hours. It is then placed in a refrigerator for holding overnight. The following day, the whole blood is processed into serum. This process includes centrifuging the whole blood at 5,000 RPM’s for 15 minutes. The serum is poured off the clot into a clean tube and centrifuged again at 5,000 RPM’s, this time for 5 minutes. The final serum is poured into a vial which has the appropriate label attached. A stopper is placed on the vial and then a crimp seal is attached. The vial of serum is then placed in a freezer, which is maintained at below -20oC. It is held in that freezer until it is packed and shipped below freezing with ice packs.