Isolation and characterization of individual functionally reactive cytotoxic T lymphocytes: conjugation, killing and recycling at the single cell level

Abstract Isolation and characterization of individual functionally reactive cytotoxic T lymphocytes have been achieved. Peritoneal exudate cytotoxic lymphocytes were obtained from BALB/c mice injected with EL4 tumor cells. Lymphocyte tumor cell conjugation was promoted by centrifugation. Individual conjugates comprised of one lymphocyte bound to one tumor cell were isolated with a micropipette. The ultrastructure of isolated killer lymphocytes and the lysis of conjugated target cells were analyzed. The cytotoxic lymphocytes are small cells with an indented nucleus which is poor in peripheral chromatin and rich in rough nuclear sap. The cytoplasm contains one‐membrane‐bound lysosome‐like granules and clusters of ribosomes, but no rough endoplasmatic reticulum. The Golgi apparatus is well developed. Direct evidence obtained at the single cell level shows that a single effector lymphocyte is required and sufficient for the destruction of a single target cell and that killer cells which have been responsible for the lysis of a given target cell can lyse a second and even a third time.