Detection of a novel 40 000 MW excretory Toxoplasma gondii antigen by murine Th1 clone which induces toxoplasmacidal activity when exposed to infected macrophages

To analyse target molecules of the CD4+ T‐cell response to toxoplasma infection, a panel of Toxoplasma gondii‐specific murine CD4+ T‐cell clones has been established. Clone 3Tx15, belonging to the T helper 1 (Th1) subtype, abolished intracellular parasite growth when co‐cultured with macrophages and live toxoplasma at a ratio of 2:2:1. This effect results from macrophage toxoplasmicidal activity induced upon parasite‐dependent cellular interaction, an irrelevant Th1 clone failed in this three‐party system. Clone 3Tx15 detects its corresponding antigen in the supernatant of infected cells and also reacts with a host cell‐free preparation of T. gondii‐excreted/secreted antigens. T‐cell blot analysis of two‐dimensionally separated toxoplasma lysate revealed a molecular weight of about 40 000 for the fractions stimulating clone 3Tx15. As checked in parallel enzyme‐linked immunosorbent assay, the 40 000 MW T‐cell antigen co‐migrates with the excretory protein GRA4, the sole 40 000 MW T. gondii antigen hitherto known to be recognized by T lymphocytes. Nevertheless, neither recombinant GRA4 nor immunoaffinity‐purified natural GRA4 was stimulatory for clone 3Tx15. Our findings thus demonstrate that Th1 clone 3Tx15 which induces toxoplasmicidal activity during antigenic interaction with infected macrophages defines a new 40 000 MW excretory T. gondii antigen.