A murine monoclonal antibody (MAb) directed against a surface-exposed epitope of the lipooligosaccharide (LOS) of Haemophilus ducreyi strain 35000 was shown to be reactive with all 37 strains of this pathogen tested in a colony blot-radioimmunoassay. The LOS epitope bound by this MAb appeared to be stably expressed by H. ducreyi growing in vitro. The use of this MAb in the immunolimulus system revealed that it could detect purified H. ducreyi LOS at a level of 25 pg/ml. Similarly, this immunolimulus system could detect as few as 1000 colony forming units of in vitro-grown H. ducreyi cells per ml of buffer. When this MAb was utilized in the immunolimulus system together with lesion material from rabbits infected with two different H. ducreyi strains, a positive reaction was obtained with every sample tested, even when no viable organisms were present in the lesion material. In contrast, this MAb yielded consistently negative results when used in the immunolimulus system with lesion material from animals infected with Staphylococcus aureus.