We used an antibody-capture enzyme immunoassay (EIA) to evaluate the early antibody responses to Borrelia burgdorferi in paired sera from 30 patients with erythema chronicum migrans. During acute disease, 20 (67%) patients had elevated specific IgM responses, and by convalescence (one to four weeks after treatment), 28 (93%) patients had increased IgM or IgG responses. In acute specimens, elevated IgM responses correlated with disseminated infection; however, by convalescence, most patients with either localized or disseminated disease had positive tests. Among 133 control subjects, IgM cross-reactivity was observed in 4 of 37 patients with either Epstein-Barr virus or rickettsial infections, and false-positive IgG tests were seen in 8 of 28 patients with syphilis. With antibody-capture EIA, the diagnosis of Lyme disease can be confirmed in the majority of acutely ill patients and in almost all patients by convalescence.