A Method for Producing Specific Antisera With Small Doses of Immunogen

We have succeeded in producing specific antisera to the subunits of HCG and testosterone conjugates with one immunization of 20 to 100 μg of immunogen. The potential of the method for producing specific antisera to glycoprotein and other hormones with limited amounts of purified material prompts this report. CR 100α and CR 100β, the subunits of HCG,. were prepared by Drs. R. E. Canfield and F. Morgan (1). Testosterone-3-(0-carboxymethy1) oxime was coupled to either bovine serum albumin (Armour) or keyhole limpet hemocyanin (Calbiochem) (2). Equal volumes of complete Freund's adjuvant, a 0.15 M NaCl solution of the immunogen and dried tubercle bacilli (2.5 mg/ml of total mixture) were homogenized for 5–10 minutes at increasing speeds to maximum setting in a Sorvall Omnimixer. Fur was shaved from the back and proximal limbs of 3 month old New Zealand white rabbits and the emulsion administered intradermally at 30–50 sites so that each animal received a total of 2 ml of the emulsion. At a separate site 0.5 ml of crude Bordetella pertussis vaccine (E. Lilly Corp.) was injected intradermally.