Rapid and precise immunotyping of Chlamydia trachomatis was achieved by testing whole organisms (elementary bodies) in the microimmunofluorescence test with monoclonal antibodies. Monoclonal antibodies were produced with standard techniques by using an immunization schedule that encouraged the development of immunotype-specific antibodies. Fifteen monotypic or multitypic (subspecies) monoclonal antibodies were chosen for use in a two-step typing system that required strains of C. trachomatis to be tested against six to eight monoclonal antibodies for classification. Immunotyping with monoclonal antibodies was studied by testing 313 strains, typed with the previous method that utilized immunized mouse antisera, that represented each of the 15 established serovars. The two-step monoclonal antibody method resulted in a classification similar to the current one. Only one strain typed differently with the two methods. With the monoclonal antibody method, available lymphogranuloma venereum (LGV) serovars L1 and L3 could not be differentiated from trachoma serovars E and G, respectively, unless the strains had been identified as LGV. Monoclonal antibody typing was simpler to perform and more precise; it allowed easy differentiation between closely related serovars. Three new types were discovered among the strains previously classified as serovars D, I, and L2. These are tentatively being considered subtypes and are labeled D', I', and L2'.