Significance The lack of commercially available HTLV seroconversion panels has made it necessary to rely on animal models to evaluate antibody development during the course of HTLV infection. These specimens are the only alternative to dilutional samples by which the sensitivity of HTLV screening tests can be judged. Objectives The objectives of this study was to the time course, and sequence of appearance of antibodies against HTLV-1 and HTLV-2-specific viral antigens in cynomolgus macaques following experimental intravenous inoculation with HTLV-I or HTLV-II infected cell lines. Weekly plasma samples from inoculated animals were tested in a variety of EIA and Immunoblot assays. Results Antibody to p19 was detected as early as week 1 by immunoblot, however, licensed EIA tests based on viral lysate only (week 3) or spiked with recombinant p21e (week 2-3) were less sensitive. No antibody to native env was observed by blots, but antibody against recombinant p21e was detected in both animals at week 2, and against recombinant gp46 was detected in one monkey at week 2 pi. An antibody capture microter plate ELISA that uses recombinant transmembrane env (p21e) and gag (p24) proteins from both HTLV-I and HTLV-II was able to detect antibody at week 1 pi. The increased sensitivity of this new assay was due solely to the p21e antigens and the assay format which can detect both IgG and IgM antibodies. Animals inoculated with HTLV-II showed no evidence of infection and remained seronegative at 16 weeks p.i. Future Directions Reinoculation studies using different HTLV-II cell lines, and at different doses, will be performed on both animals that failed to seroconvert, with the objective of generating an HTLV-II seroconversion panel. These reagents will used to evaluate commercial diagnostic assays currently in development. KEY WORDS human T-cell leukemia virus FUNDING Ortho Clinical Diagnostics