Gene-modified lymphocytes have a potential role in the therapy of cancer, infectious diseases, and genetic disorders of the immune system. Current gene therapy protocols involving gene transfer into lymphocytes utilize retroviruses with amphotropic envelope proteins. However, transduction efficiencies in lymphocytes using these viruses are relatively low. A potential strategy to improve gene transfer efficiency is the utilization of alternative retroviral envelopes that target unique receptors on the cell surface. One such alternative retroviral envelope, the gibbon ape leukemia virus (GALV) envelope, targets a distinct surface receptor (GLVR-1) that is 60% homologous but not cross-reactive to the amphotropic receptor (GLVR-2/RAM-1). Understanding the relationship between receptor expression and transduction efficiency is important for designing new strategies to improve gene transfer. Therefore, we compared GLVR-1 and GLVR-2 mRNA levels in lymphocytes and found that GLVR-1 was expressed 8- to 19-fold higher than GLVR-2. We then analyzed whether this enhanced expression of GLVR-1 correlated with increased infectivity of lymphocytes by retroviral vectors that utilize the GALV envelope compared to those that use the amphotropic envelope. We evaluated retroviral vectors packaged with either PA317 or PG13, which express the amphotropic and GALV envelopes, respectively. Lymphocyte transduction with PG13-packaged vectors was 4- to 18-fold higher than that with PA317-packaged vectors. These findings suggest that receptor expression level is an important factor in retroviral-target interactions and that gene transfer into human T lymphocytes should be performed with retroviruses that use the GALV envelope as opposed to retroviruses that use the amphotropic envelope.