The proliferative responses to four gag peptides were examined in 24 HIV-seropositive patients whose CD4 counts ranged between 500 and 1400 cells/mm 3. To overcome some of the limitations imposed by HIV infection on the T-cell proliferative assay, recombinant interleukin 2 (rIL-2) was added to the cultures, and the culture time of the cells was increased from the standard 6 to 8 or 10 days. Four of 24 patients responded to one or more core peptides, aa 180-194, 208-217, 267-286, and 287-306 by the standard 6-day culture: this increased to 13 of 24 using the optimized culture approach. The greatest number and magnitude of responses occurred after cells were in culture for 8 days. Eight patients responded to gag 180-194, which has not been identified previously as a TH epitope in humans but has considerable homology with a TH epitope recognized by cloned T cells from macaques immunized with simian immunodeficiency virus (SIV). We have identified four T-cell epitopes on the HIV core protein p24, using synthetic peptides as immunogens. Three of the peptides would not have been considered immunogenic had the standard assay system been used to detect T-cell responsiveness. We have also shown that a region of the core protein encompassed by aa 180-194 is recognized by TH cells in humans.