Protein antigens of Plasmodium knowlesi clones of different variant antigen phenotype

Mature asexual stages of the malaria parasite Plasmodium Knowlesi synthesize proteins of Mr 180 000-225 000 that are expressed on the outer membrane of infected erythrocytes and which vary antigenically such that different parasite clones are specifically agglutinated with homologous antibody. Other non-agglutinable clones have been prepared which fail to express variant antigen on infected cells. Two agglutinable clones of different variant antigen phenotypes and a non-agglutinable cone were examined to determine the proportion of total malarial proteins represented by variant antigens. Malarial proteins were labelled with various radioactive amino acids and the sodium dodecyl sulphate--polyacrylamide gel patterns for the three clones compared by fluorography. The patterns were indistinguishable, the variant antigens being undetectable in analyses of total malarial proteins. Furthermore, these antigens were not detected by Coomassie Blue-staining of total cellular proteins after electrophoresis. Sodium dodecyl sulphate and Triton X-100 extracts of labelled cells were immunoprecipitated using a panel of sera of defined agglutination specificity. The variant antigens could not be detected in the fluorographic patterns of total malarial antigens immunoprecipitated by these sera. In contrast, after lactoperoxidase catalysed radio-iodination of intact schizont-infected cells, the 125I-variant antigens on the cell surface were identified by demonstrating their accessibility both to antibody and to trypsin with intact cells. Thus, the variant antigens are quantitatively very minor malarial proteins that can only be detected by methods which selectively analyse the subset of proteins on the erythrocyte surface.