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In vitro induction of cell mediated immunity to murine leukemia cells part 2 cyto toxic activity in vitro and tumor neutralizing capacity in vivo of anti leukemia cyto toxic lymphocytes generated in macro cultures



In vitro induction of cell mediated immunity to murine leukemia cells part 2 cyto toxic activity in vitro and tumor neutralizing capacity in vivo of anti leukemia cyto toxic lymphocytes generated in macro cultures



Journal of Immunological Methods 16(1): 39-58



Highly reactive cytotoxic lymphocytes (CTL) against syngeneic and allogeneic murine leukemia cells were generated in vitro in macro 1-way mixed leukocyte-tumor cultures (MLTC). Cultures set up in large tissue culture flasks contained up to 400 .times. 106 normal spleen cells (responder cells) and 20-40 .times. 106 mitomycin C-treated leukemia cells (stimulator cells). Successful sensitization in macrocultures was greatly dependent upon the responder cell density and the responder/stimulator cell ratio. Cytotoxic activity, as measured by the 51Cr-release assay, peaked on day 5-7. Sensitized memory lymphocytes produced in primary MLTC could be restimulated with the original tumor cells to give a more rapid and stronger secondary cytotoxic response. Lymphocytes sensitized to allogenic leukemia cells reacted equally well with the sensitizing leukemia cells and with the corresponding normal lymphoid target cells, whereas lymphocytes sensitized to syngeneic leukemia cells did not react with the homologous normal lymphocytes. Cryopreserved normal splenocytes and leukemia cells were as efficient as fresh cells in generating allogeneic and syngeneic CTL. Using a Winn-type tumor neutralization assay, it was shown that allogeneic and syngeneic splenocytes sensitized in vitro to EL4 leukemia (of C57BL/6 mice) and to YAC leukemia (of A mice) were capable of preventing tumor growth in the syngeneic host, whereas cultured normal splenocytes frequently showed a tumor-enhancing effect. Long-term survivors, remaining after inoculation of leukemia cells and sensitized lymphocytes, also became resistant to a tumor challenge that was up to 10,000 greater than the minimum lethal dose.

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