Inhibition of clone formation as an assay for T cell-mediated cytotoxicity: short-term kinetics and comparison with 51Cr release.
The short-term kinetics of T cell-mediated cytotoxicity was investigated using a cloning inhibition assay. Murine cytotoxic thymus-derived lymphocytes (CTL) generated in vitro in mixed leukocyte cultures (MLC), were incubated for various periods of time at 37 degrees C with allogeneic mastocytoma target cells. The mixtures were then plated in soft agar, and mastocytoma clone formation was assessed after 5-7 days incubation. Using this technique, it was demonstrated that events leading to the loss of cloning ability could be detected after 1-3 min incubation at 37 degrees C, and after 20-30 min, 95% of the clone forming cells had been inactivated. When these results were compared directly with those obtained using the conventional 51Cr-release assay, it was found that the events leading to loss of cloning ability occurred more rapidly than indicated by the isotope assay. However, a modification of the 51Cr-release assay involving EDTA addition, gave comparable results to the cloning inhibition assay. These results raise the possibility that the events leading to 51Cr-release of tumor target cells may be related in time to those leading to the loss of cloning ability.
D000818:Animals; D002860:Chromium Radioisotopes; D002999:Clone Cells / Q000276:immunology; D003601:Cytotoxicity Tests, Immunologic; D004306:Dose-Response Relationship, Immunologic; D004492:Edetic Acid / Q000378:metabolism; D007700:Kinetics; D012515:Mast-Cell Sarcoma / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D013601:T-Lymphocytes / Q000276:immunology*
D002860:Chromium Radioisotopes; D004492:Edetic Acid