Early detection and specificity analysis of human cytolytic T lymphocyte (CTL) colonies generated in soft agarose culture: a potential assay for definition of CTL defined (CD) determinants.

In this communication we describe and early, large-scale screening assay for the detection of colonies with varied cytolytic specificity. The colonies are generated in soft agarose culture from day 3 mixed lymphocyte culture (MLC) alloactivated cells. A cell mediated lympholysis (CML) assay utilizing as few as 500 target cells has made it possible to prescreen for cytolytic T lymphocyte (CTL) colonies and to test for antigen specificities as early as 11 and 14 days, respectively, after MLC priming. Large numbers of colonies, from 80 to over 150, have been prescreened against a specific sensitizing target cell, and as many as 30 CTL colonies have been simultaneously tested against a panel of multiple targets carrying defined HLA-A, -B, -C, -DR antigens to evaluate antigen specificity. All CTL colonies are lytic against the specific sensitizing target cell and do not lyse the target autologous to the responder. Some are found to be operationally specific in that they lyse only those target cells which share HLA serologically defined (SD) antigens with the sensitizing cells, and other show cytolytic patterns which are not correlated with known HLA-SD antigens. These observations support, at a much finer level of analysis, the possible distinction between SD and CTL defined (CD) determinants.