The adult T-cell leukemia (ATL)-associated antigen complex (ATLA) is recognized by serum antibodies of carriers of ATL virus (ATLV). ATLA consists mainly of ATLV polypeptides and their precursors. The sera from 22 ATL patients, 21 healthy carriers and 9 healthy individuals were examined quantitatively by immunofluorescence assay (IF) for ATLA and by a newly developed radioimmunoprecipitation test with purified 125I-gp68, the putative env gene product of ATLV. More qualitative results were obtained by analysis on polyacrylamide gel (PAGE) of immunoprecipitates from lysates of 35S-cysteine-labelled cells producing ATLV, pelleted ATLV and cell-free culture supernatant. The two quantitative assays gave negative results with sera from all normal subjects and a few patients, but detected ATLA antibodies in all the healthy ATLV carriers. An important finding was that sera of patients that gave negative results in one assay gave positive results in the other, and vice versa. In contrast, all sera from ATL patients and healthy carriers, but not normal donors, precipitated ATLV-specific glycopolypeptides, gp68 and gp46 from 35S-labelled materials. But core polypeptides p28, p24, p19 and p15 were precipitated only by sera with IF titers of over 80. Thus, anti-ATLA antibodies in seropositive sera are predominantly directed against glycopolypeptides of ATLV, and the antibody reactivity to ATLA antigens does not differentiate between ATL patients at various stages of the disease and healthy ATLV carriers.