Excretory-secretory antigen of Mycobacterium tuberculosis H37Ra (Mtb ES antigen) isolated from culture filtrate was partially purified by 6% trichloroacetic acid precipitation. The TCA supernatant fraction (Mtb EST antigen) was examined for its diagnostic use in the detection of tuberculous IgG antibody in human sera by stick Indirect ELISA. Using Mtb EST antigen, tuberculous IgG antibody was detected in 90% of smear positive and 71% of smear negative pulmonary tuberculosis cases and 10% of healthy and disease controls. Further fractionation of Mtb EST antigen by SDS-PAGE yielded four active antigenic fractions viz., Mtb EST-3,4,6 and 10. Diagnostic evaluation of these fractions showed Mtb EST-6 antigen fraction to be useful in detection of both smear positive and smear negative pulmonary tuberculosis cases with sensitivities of 94% and 78% respectively and specificity of 88%.