Standard Immunization of Rabbits.

Rabbits can be immunized by administering biweekly injections of a purified antigen, cultured cells, or cDNA. The minimum amount of antigen capable of inducing a response will depend on the nature of the antigen and on the host, but for rabbits, the minimum dose will be in the range of 10 μg per injection, although 100 μg per injection will be used more commonly. If a pure, soluble protein antigen is being used and is abundant, then a dose of 0.5-1 mg in adjuvant at each immunization is a sensible general recommendation. The injection sites on the rabbit are shaved and disinfected before immunization. Adjuvants are mixed with the immunizing antigen for the first two immunizations only, and Complete Freund's adjuvant is only used with the first immunization; subsequent immunizations are performed in phosphate-buffered saline (PBS) or normal saline, with or without Incomplete Freund's adjuvant. Once a good titer has developed against the antigen of interest, regular boosts and bleeds are performed to collect the maximum amount of serum. For rabbits, boosts should be spaced every 6 wk, and serum samples of 20-40 mL should be collected ∼10-12 d after each boost; typically, a single sample bleed from a rabbit will yield 25 mL of serum.