Affinity separation and partial characterization of serologically active Leishmania donovani antigens

Leishmania donovani-soluble antigens capable of antibody production in rabbits were separated from the total antigenic make-up of the parasite by adsorption on to anti-Leishmania immunoglobulins coupled to CNBr-activated Sepharose 4B (ALIG). Immune rabbit sera were produced either by intravenous inoculation of living promastigotes or by intradermal and subcutaneous inoculation of soluble antigens. Approximately four times as much soluble antigen was bound to ALIG produced by i.v. than i.d. and s.c. inoculation although the percentage recovery was less. The antigenic fractions recovered were subjected to immunoelectrophoresis, isoelectric point and molecular weight determination. The results of affinity chromatography indicate high antigen-antibody interaction affinity, the recovery of at least 28 antigens (with 0.15 M NaCl, pH 11.0) and complete loss of serological activity (determined by immunoelectrophoresis) on treatment with 8 M urea.