Immunological detection of Plasmodium knowlesi antigens after electrophoretic transfer from SDS-polyacrylamide gels to diazophenylthioether paper.
Freeze-thawed extracts of mature schizonts of Plasmodium knowlesi (strain W1) were electrophoresed in sodium dodecylsulphate (SDS) polyacrylamide gels, then transferred electrophoretically to diazophenylthioether paper. The transferred antigen was probed using a purified polyvalent, polyclonal immune serum pool isolated from rhesus monkeys (Macaca mulatta). This method for identifying parasite antigens was compared with immunoabsorption, using parasites which had been metabolically labelled in vitro with 35S-methionine. The methods showed general correspondence but some antigens failed to react after transfer because of loss of antigenicity during SDS-polyacrylamide gel electrophoresis (SDS-PAGE). Some antigens were observed preferentially after the transfer procedure, possibly because of their failure to elute from the immunoabsorbant or because they may have a low methionine content. The transfer method failed to reveal the antigen reactivity of three out of four monoclonal antibodies tested, apparently because the epitopes involved were inactivated by SDS. During P. knowlesi infection in the resistant kra monkey (Macaca fascicularis) the transfer method showed the presence by day 10 of antibodies against two components having an apparent molecular weight by SDS-PAGE similar to those of putative protective antigens described in other studies using monoclonal antibodies.
D000818:Animals; D000911:Antibodies, Monoclonal; D000941:Antigens / Q000302:isolation & purification*; D003979:Diazonium Compounds; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D007074:Immunoglobulin G / Q000096:biosynthesis; D007075:Immunoglobulin M / Q000096:biosynthesis; D008252:Macaca fascicularis; D008253:Macaca mulatta; D008288:Malaria / Q000276:immunology; D010961:Plasmodium / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000941:Antigens; D003979:Diazonium Compounds; D000939:Epitopes; D007074:Immunoglobulin G; D007075:Immunoglobulin M; C030003:diazophenylthioether