Development of graft-vs.-host disease-like syndrome in cyclosporine- treated rats after syngeneic bone marrow transplantation. I. Development of cytotoxic T lymphocytes with apparent polyclonal anti-Ia specificity, including autoreactivity

Cell-mediated Lympholysis Assay (CML). The CML assay was performed using standard techniques as previously described (1 i, 17) . Briefly, spleen ceils from the test animal, fractionated over nylon wool to enrich for T cells (18) , were used as effector cells in the CML assay. Target cells were either 3-d-old phytohemagglutinin (PHA)-induced blast cells or 4-d-old concanavalin A (Con A)-induced blast cells labelled with 400/~Ci of ~Cr. Maximum, and spontaneous ~lCr release were obtained by adding 0.1 ml 1 N HCI or 0.1 ml of complete medium, respectively, to wells containing 104 target cells. Percent specific ~TCr release was calculated according to the following formula: % specific 5~Cr release --(cpm experimental -cpm spontaneous) x 100/(cpm maximum -cpm spontaneous). Effector/target ratios, in most instances, for these assays, were 100:1.