Corynebacterium ulcerans strain ATCC 9015 develops during growth in environmental media an exotoxin which is lethal for guinea-pigs. The exotoxin has been purified by ion-exchange chromatography on DEAE-Sephadex A-50, affinity chromatography using immunosorbent techniques, and gel-filtration through Sephadex G-75. The exotoxin is of a protein nature and consists perhaps of two subunits. The molecular weights of the exotoxin and its subunits as estimated by SDS-PAGE were 40 000, 25 000, and 15 000, respectively. The toxic properties of exotoxin were destroyed upon heating (60 degrees C, 30 min), treatment with trypsin (10 micrograms/ml, 37 degrees C, 30 min), and by the extremes of pH (pH 4.0 or 10;0 24 hrs). Homologous antibodies readily neutralized the exotoxin while commercial diphtheria antitoxin had no effects. No immunological cross-reactivity was revealed between exotoxin of C. ulcerans and other bacterial toxins such as diphtheria toxin, enterotoxin from V. cholerae, and exotoxin A of P. aeruginosa.