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Biological assay for Clostridium perfringens enterotoxin with Vero cells.

A simple and reproducible assay method using African green-monkey kidney (Vero) cells for Clostridium perfringens enterotoxin was developed. Vero cells were incubated with enterotoxin in wells on a microtiter plate for 60 min at 37 C and stained with a 0.2% nigrosin solution. A nearly linear curve was obtained between the ratio of the stained cells and the log dose of purified enterotoxin ranging from 25 to 400 ng/ml. As culture supernatant of some strains in Duncan and Strong (DS) medium caused false positive reaction, the proposed method may not be reliable for assaying enterotoxin in culture at a concentration of 250 ng/ml or lower. Dilution of the culture supernatant by 10-fold or higher with buffered saline eliminated such false positive reaction. The Vero-cell staining assay, therefore, is applicable for assaying enterotoxin in pure form and that in cultures in DS medium at 250 ng/ml or higher concentrations. It seems essential to prove that the reaction is neutrarized with anti-enterotoxin serum.