Antigenic components of the protozoan parasite Leishmania enriettii inducing humoral and cell-mediated immune responses in infected guinea-pig

Abstract

Subcellular fractions of Leishmania enriettii promastigotes were prepared by differential centrifugation of parasite homogenates. Antigenic activities of these fractions were determined by microcomplement fixation and indirect radioimmunoassay methods using L. enriettii immune guinea-pig serum. Results showed that the membrane fraction had major antigenic activity although other fractions also inhibited some activity. This distribution of antigenic activity in different fractions was similar to that of 5′-nucleotidase, a cell surface marker enzyme. Attempts to radiolabel L. enriettii promastigotes by lactoperoxidase catalyzed iodination technique achieved very limited success. A highly active antigenic fraction was prepared from freeze-dried parasites by differential and density gradient centrifugation. This fraction was about 200 times more active than intact parasites and contained a considerable amount of polysaccharides along with some proteins and lipids. Further analyses showed that the polysaccharides were largely involved in the antigenic activity of this material with immune guinea-pig serum. Antigenic activities of different L. enriettii fractions were also tested in cell-mediated reactions by skin testing of immune guinea-pigs and in vitro transformation of splenic lymphocytes as measured by [ 3H]thymidine uptake. A high mol. wt fraction (ribosomal fraction), prepared by passing the supernatant through a Sephadex G-200 column, was rather potent in these reactions. Interestingly, delayed hypersensitivity reactions could be substantially eliminated by pronase treatment of the fraction suggesting the protein nature of the antigenic determinants. These results indicate the heterogeneity of L. enriettii antigens and suggest that major humoral and cell-mediated components of the immune response in infected guinea-pig are directed against different antigenic determinants of the parasite.