Murine anti-Treponema pallidum monoclonal antibodies were employed in the detection of low numbers of pathogenic treponemes. Monoclonal antibodies were used as a primary antibody source in a solid-phase immunoblot assay system. All monoclonal antibodies assayed were capable of detecting ca. 1.0x103 to 2.5x103 treponemes. Of 13 monoclonal antibodies examined, 3 were able to detect 103 virulent treponemes, and 1 of these antibodies was able to reveal the presence of as few as 500 organisms. Western blot analyses showed that all anti-T. pallidum monoclonal antibodies exhibiting high sensitivities for the detection of T. pallidum cells were directed against an abundant, 47,000-48,000 dalton surface-exposed antigen of the organism. With two possible exceptions, the monoclonal antibodies tested reacted specifically with T. pallidum, either purified or found within a high-contaminating tissue background, and not with Treponema phagedenis biotype Reiter, Haemophilus ducreyi, Neisseria gonorrhoeae, herpes simplex virus type 2, or normal rabbit testicular tissue.