Clostridium difficile toxin is frequently encountered in patients with antibiotic-associated diarrhea. A commercially available enzyme immunoassay (EIA) detecting toxins A and B was evaluated, screening 148 stool specimens specifically submitted for the detection of C. difficile. A sensitivity of 100% and a specificity of 97.5% were found compared to a tissue culture assay. The overall prevalence of C. difficile toxin was 22.1%. Cephalosporins had been administered significantly more often to toxin-positive patients. The EIA proved to be a suitable test for the detection of C. difficile toxin, revealing high prevalences in specifically submitted clinical specimens.