T cell subsets defined by expression of Lyt-1,2,3 and Thy-1 antigens. Two-parameter immunofluorescence and cytotoxicity analysis with monoclonal antibodies modifies current views.

Abstract

Antibodies to mouse T cell antigens such as Thy-1, Lyt-1, Lyt-2, and Lyt-3 have been used to characterize T cell subpopulations and to determine precursor-progeny relationships (1). Although the original Lyt-l+2 ÷, Lyt-l+2 -, and Lyt-l-2 ÷ classifications have been very useful, several recent findings appear to be inconsistent with the model proposed by Cantor and Boyse, i.e., that Lyt-l+2 ÷ immature T cells give rise to Lyt-1+2-helper cells and Lyt-1-2 + cytotoxic and suppressor cells (1) For example, cytotoxic T cells originally classified as Lyt-1-(2) have in some cases been depleted by anti-Lyt-1 and complement (3-5). Furthermore immunofluorescence studies in thymus (6) and peripheral lymphoid tissue (7, 8) have indicated that Lyt-1 is expressed on a larger proportion of cells than would be expected from the cytotoxic depletion studies (2). The immunofluorescence studies (5, 7) also showed that Lyt-l+2 -cells comprise -50% of the cortisone-resistant thymocytes, indicating that T cells are already divided into separate lineages in the thymus.Quantitative examination of the question of Lyt-1, Lyt-2, and Lyt-3 expression has recently been facilitated by the production of monoclonal rat antibodies to Thy-I and Lyt antigens (7). These antibodies can be purified to a single molecular species and thus can be used at high antibody/cell ratios in cytotoxic assays. They also can be directly conjugated with fluorochromes and be used with very low backgrounds in quantitative single-cell fluorescence assays for measuring determinant expression by flow cytometry with the fluorescence-activated cell sorter (FACS) We show here that although subpopulations can be clearly distinguished on the basis of quantitative phenotypes for these antigens, the qualitative designations Lyt-1-2 + and Lyt-l+2 + need to be modified. Lyt-l+2 -cells express higher levels of Lyt-I than do Lyt-1 +2 ÷ cells, and are also more easily killed by anti-Lyt-1 plus complement We also show that although all Thy-1 ÷ cells also bear Lyt-1, there is a clear negative correlation in the surface density of these two antigens. There is a small subpopulation

Differentiation antigens identify subpopulations of rabbit T and B lymphocytes. Definition by flow cytometry.

“…Both human and mouse splenic T populations contain two antigenically and functionally distinct subsets: helper-inducer and suppressor-cytotoxic, which are specifically recognized by a series of monoclonal antibodies (3)(4)(5)(6). In terms of percentages of cells stained and staining profiles, none of the anti-rabbit monoclonals closely resembled the subset-specific anti-mouse or anti-human reagents.…”

Precursor frequency analysis of lymphokine-secreting alloreactive T lymphocytes. Dissociation of subsets producing interleukin 2, macrophage-activating factor, and granulocyte-macrophage colony-stimulating factor on the basis of Lyt-2 phenotype.

“…By the combination of flow cytometry, a microculture system for the activation of unprimed lymphocytes at limiting dilution, and sensitive microassays for cytolytic and lymphokine activities, it has been possible to compare directly in each series of microcultures the frequencies of lymphokine-secreting cells and cytolytic T lymphocytes in populations separated according to Lyt-2 phenotype. The Lyt-1 phenotype was not taken into consideration in this study in view of recent demonstrations by several laboratories that this marker is expressed by essentially all T lymphocytes (24)(25)(26).…”

The secondary in vitro murine cytotoxic T cell response to the flavivirus, West Nile

“…Both Pang et al (27) and Ashman and MuUbacher (28) have demonstrated that for Lyt 2+ cytotoxic T cells to be generated in vitro, Lyt 1 ^ cells are required to be present. In this work we used the L3T4 surface antigen as a marker for helper T cells rather than the Lyt 1 antigen because expression of L3T4 and Lyt 2 by spleen cells appears to be mutually exclusive (29) and the expression of L3T4 differs from that of Lyt 1 in that Lyt 1 is expressed by all splenic T cells, albeit at a lower density on Lyt 2 cells (30).…”