Analysis of immune responses involving distinctive cell types has been greatly facilitated by the development of technology for separating and purifying different lymphocyte populations (1–5). In conjugation with the use of markers which identify selective expression of cell surface components (6, 7) this approach provides an efficient means of studying multi-component immune responses. Marker systems identifying human thymus derived 2 (T) and bone marrow-derived (B) lymphocytes are now well advanced (reviewed in 8–10) and we report here the application of various markers to determine the efficacy of separation techniques aimed at providing a simple and reproducible protocol for the preparation of effectively pure human T and B lymphocytes for functional studies. Materials and Methods. All cell separations were performed on human tonsil cell suspensions since these were available daily from University College Hospital ENT Theatre and provided a convenient source of over 2 × 108 viable lymphocytes per tonsil.