Abstract
Sixty-three homozygous rabbits from five different partially inbred strains were hyperimmunized with either type III or VIII pneumococcal vaccine for 1–3 courses of immunization, Fifty-six percent of the ACEP strain rabbits and 46 per cent of the New Zealand White type III strain demonstrated the presence of either electrophoretically restricted or partially restricted antibody molecules during the primary or secondary courses of immunization. None of the type III c, AX or ACCR(B) strains of rabbits gave either restricted responses when immunized with the pneumococcal vaccines. Three of the ACEP strain rabbits and two of the type III strain produced large quantities of highly restricted antibodies and were used to initiate structural studies on the antigen-combining site of antibody molecules. The rabbit antibodies were purified initially by affinity chromatography. The anti-SVIII antibodies were all eluted in a single fraction from an SVIII immunoadsorbent. The anti-SIII antibodies were all eluted in two fractions (one in various aqueous buffers and the other in guanidine-HCl) from an SIII immunoadsorbent. The specificially purified antibodies were examined analytically by isoelectric focusing in polyacrylamide gels and then were further fractionated by preparative liquid isoelectric focusing. Each purified antibody preparation from a restricted responder yielded 2–4 major electrofocused antibody fractions. The isoelectric point difference between the first and last eluted focused antibody fractions varied between 0·2–0·8 pH unit. The affinity chromatography and the isoelectric focusing procedures provide a successful means of preparatively obtaining different antibody fractions even from a single highly restricted antiserum.
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