A micromethod for prospectively testing lymphocyte-mediated cytotoxicity (LMC) using fresh or frozen cells labelled...

A direct, specific and rapid method has been developed to prospectively evaluate lymphocyte mediated cytotoxicity (LMC) for kidney transplant recipients. Donor lymphocytes, labeled with fluorescein diacetate, were incubated in a microtest plate for 2 h with recipients' effector cells. The percentage of fluorescent cells in each well was estimated relative to controls using an inverted phase fluorescence microscope. Lymphocytes obtained from seven of 12 recipients post-transplantation exhibited positive reactions, fluorescence of labeled donor cells was reduced to 30-65% of that observed in controls, and these reactive patients required nephrectomies. The responses of the remaining five patients were negative. Tests with their donor cells showed 80-95% control fluorescence, with favorable transplant results. Six of 21 recipients evaluated prospectively required nephrectomies within 34 days post-transplantation. Three of these subjects showed 15-65% control fluorescence, a positive LMC response; the other three yielded negative LMC responses but required nephrectomies possibly due to renal ischemia prior to transplantation. The remaining 15 patients demonstrated 85-90% control fluorescence with graft function continuing for 17 to 270 days. The results from direct LMC tests with a fluorescent label indicate that the system is of value in predicting early rejection of renal allotransplants.