Interaction of Clostridium difficile toxin A with L cells in culture

Toxin A of Clostridium difficile was purified by column chromatography and acetic acid precipitation. Cells exposed to toxin A showed polarization of nuclei towards one pole of the cells. Toxin A was conjugated to ferritin and applied to L cells to localize binding sites of this toxin to the cell surface. It was found that toxin A conjugate attached to the cell membrane in aggregated form. Antibody specific to toxin A was prepared and used for localization of intracellular toxins in intoxicated cells. Toxin A was found inside the cytoplasm 6 h after cell treatment, mainly in the form of aggregates inside the cytoplasmic vacuoles. At 24 h after exposure, toxin A could be detected within the cytoplasm. Tunicamycin treatment of cells reduced the cell-binding efficiency of toxin A to 50%, but neuraminidase did not effect toxin binding significantly.