Ultrastructural localization of an intracellular Toxoplasma protein that induces protection in mice

Abstract

We report the ultrastructural localization of Toxoplasma antigens recognized by monoclonal antibody F3G3, which protects mice against lethal challenge. By using colloidal immunogold labeling, F3G3 failed to react with the surface of intact Toxoplasma cells, confirming previous observations that it recognizes an intracellular antigen. Immunoperoxidase labeling with F3G3 was obtained only when Toxoplasma cells were previously exposed to Triton X-100. A specific immunoperoxidase reaction was located beneath the surface membrane in the region of the pellicle and within the elaborate network of vesicles which are extruded from the surface of Toxoplasma cells during entry into host cells. Similar results were obtained with mouse polyclonal sera and a second monoclonal antibody, 6-86-1E11, both of which were produced against the F3G3 affinity-purified protein. The location of the epitope recognized by F3G3 was confirmed by purifying T. gondii-derived surface membrane vesicles, separating these proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and immunoblotting. Monoclonal antibody 6-86-1E11 and the analogous polyclonal sera reacted with the antigenically related proteins of 58 and 28 kilodaltons that were found both in the intact organisms and in the purified surface membrane vesicles. These results indicate that the epitope recognized by F3G3 is located beneath the Toxoplasma cell surface membrane and is contained within plasma membrane-derived vesicles.