Although bovine leukemia virus (BLV) is mainly associated with infections of B-lymphocytes, we have previously reported the statistically significant increase in the T-lymphocytes obtained from BLV-infected asymptomatic aleukemic (AL) cattle. In this report the presence of BLV provirus in the DNA of immunoaffinity purified T-lymphocytes from AL animals was assessed using a highly specific radiolabelled (32P) BLV-DNA provirus probe and solid phase DNA hybridization. The BLV provirus was found in the DNA of the peripheral blood mononuclear cells of all AL animals tested and three of the four purified T-lymphocyte preparations from these animals. The purified T-lymphocyte preparations used in this study contained less than 4% detectable B-lymphocytes. One animal had no detectable B-lymphocytes in the purified T-lymphocyte preparation and the DNA from these cells also gave positive hybridization results. The lymphocyte blastogenesis assay was then used as an indicator of the functional ability of lymphocytes from these BLV-infected AL cattle to respond to mitogenic stimuli. The responsiveness of lymphocytes from these animals to the mitogens concanavalin A (Con A), phytohemagglutinin (PHA), and pokeweek mitogen (PWM) was comparable to that of lymphocytes from BLV-negative animals when changes in 3H-thymidine uptake (c.p.m.) were used as measurement of mitogenic-induced blastogenesis. This indicated that infection of the T-lymphocytes by BLV does not appear to alter the overall response of the lymphocyte populations to mitogenic stimuli. High levels of spontaneous blastogenesis in the absence of mitogenic stimulation were observed for lymphocyte preparations of AL animals. The reason for this proliferation of lymphocytes is unclear; however, sera from these AL animals were found to contain a blastogenesis-augmenting factor(s) when added to lymphocytes from BLV-negative control animals in the presence of Con A, PHA and PWM.