Bovine peripheral blood lymphocytes (PBL's) from 3 cows and 1 steer infected with bovine leukemia virus (BLV) were separated by fractionation through nylon wool columns into nylon-adherent and nonadherent cell populations. Nylon-adherent cells were enriched in B-lymphocytes, as determined by the presence of surface membrane immunoglobulins (slg), whereas nylon-nonadherent cells or “non-B-lymphocytes” contained few slg-bearing cells. PBL’s and separated B-and non-B-lymphocyte populations were assayed for the presence of BLV by the induction of syncytia in bovine embryonic spleen cells. PBL’s and B-lymphocyte populations both produced many syncytia, whereas non-B-lymphocytes yielded few or no syncytia. The specificity of syncytia induction for BLV was demonstrated by neutralization of syncytia formation by anti-BLV serum. PBL’s from 2 control animals were negative for syncytia induction. This study presents further evidence that B-lymphocytes are the target cells for BLV infection.