The Chlamydia trachomatis immunodominant major outer membrane protein (MOMP) is both a target of neutralizing antibodies and the serotyping antigen and thus has been a focus of diagnostic, seroepidemiologic, and experimental investigations. The microimmunofluorescence (MIF) test has been the principal tool in serologic investigations of chlamydial infections but is difficult and expensive for routine use; moreover, since it uses whole organisms as antigen, it is incapable of revealing the molecular specificity of the humoral response to infection. These limitations were resolved by using synthetic peptides corresponding to serovar-specific antigenic regions of MOMP in an ELISA-based format to analyze the serospecificity of sera from trachoma cases. The ELISA reaction to the surface-exposed MOMP sequence variable segment 1 was immunodominant and serovar-specific and was in concordance with serovar specificity according to paired MIF test determinations. Understanding the patterns of humoral responses to MOMP determinants in patient populations will advance our knowledge of their role in the immunobiology of naturally acquired infection.