The enzyme-linked immunosorbent assay (ELISA) is a sensitive and specific serodiagnostic method for leishmaniasis. In this report, we describe how this versatile assay can be improved by the use of protein A or protein G conjugates for the specific detection of Leishmania antibody in the sera of patients with visceral leishmaniasis. In direct comparisons with anti-immunoglobulin conjugate, enzyme-linked protein A gave significantly higher absorbance values for positive sera without a corresponding increase in absorbance values for sera from normal individuals or from patients with other diseases known to cross-react with leishmaniasis. The effect was to increase the distance between positive and negative values, which aided in the interpretation of the results. This also permitted visual distinction between positive sera and negative or weakly reactive sera. The assay was effective using either blood or serum as the source of primary antibody. A further advantage of protein A over anti-Ig conjugate was its ability to detect specific antibody in dog as well as human sera. Finally, we demonstrated the usefulness of the protein A ELISA with a recombinant leishmania antigen, gp63.