Lymphocyte-mediated cytotoxicity microassay by cytofluorography

A cytofluorometric method for measuring lymphocyte-mediated cytotoxicity is described which employs acridine orange and ethidium bromide stains. These stains permit the rapid (approximately 45 sec per sample) differential enumeration of viable cells and are therefore valuable in the preparation and performance of a cytotoxicity assay. Reproducibility of viable target cell counts is good and, statistically, the same mean result is obtained when the cytofluorometric and an accepted radioisotopic method are compared. It is found that the method employing radioisotopic label ([125I]iododeoxyuridine) non-specifically killed 80% of the target cells, whereas the cytofluorographic method does not result in non-specific kill. Therefore, the cytofluorometric method reflects in vivo conditions more accurately. The cytofluorometric method also provides a more rapid and less technically demanding assay than those previously described. (Auth.)