We investigated antibodies against pX gene product, p40tax, by ELISA using recombinant p40tax protein in HTLV-I seropositive carriers as well as patients with adult T cell leukemia (ATL) and HTLV-I-associated myelopathy (HAM). Seventy (49.0%) out of 143 HTLV-I healthy carriers were found to be positive for antibody against p40tax antigen and the follow-up samples at two-year intervals revealed constant reactivity by ELISA in each carrier. The onset of antibody production was delayed 4 to 12 weeks as compared with anti-HTLV-I in primary infection cases. The anti-p40tax-positive rate (90%) in HAM patients was significantly higher than that of healthy carriers, acute and chronic ATL patients and their family members. Furthermore, HAM patients and a few healthy carriers showed high reactivities by ELISA. Children from mothers with anti-p40tax showed a higher anti-HTLV-I-positive rate than that of children from mothers without anti-p40tax (54.5% versus 12.5%). Two men without anti-p40tax and one female with low anti-p40tax developed ATL during follow-up studies. These results suggest that HTLV-I carriers could be divided into 2 or 3 sub-populations according to antibody response to p40tax. A smaller population with anti-p40tax, especially a high antibody reactivity, could have a high risk of developing HAM and of transmission from mother to child. In addition, ATL may occur in a population with low or absent anti-p40tax.