Clostridium perfringens phospholipase C (PLC), commonly known as alpha toxin, is the lethal, dermonecrotic toxin produced by all strains and is considered a major virulence factor in clostridial myonecrosis. We developed a capture antibody ELISA that accurately and specifically quantitates alpha toxin produced by C. perfringens. Another PLC, derived from Bacillus cereus, and culture filtrates from various bacterial species including Clostridium bifermentans and Clostridium novyi were not cross-reactive in this ELISA. Standard curves generated with homogenous C. perfringens alpha toxin revealed detection limits of 19 ng/ml. The ELISA was more sensitive in detecting alpha toxin than techniques such as PLC enzymatic activity and mouse lethality assays.