SUMMARYAn examination of some parameters of the lymphocyte cytotoxicity test was undertaken to see if the sensitivity of the technique could be improved by variations in serum and complement incubation time and in diluents, ionic strength, and pH. The results indicate that the optimum incubation times of serum and complement in the two-stage test are 15-20 and 20 min, respectively, although strongly reactive cells were positive after only 1 min of serum incubation. Using barbital buffer, 5% bovine serum albumin in saline, 2% dextran in saline, Plasmagel, 5% human serum, or heat-inactivated normal rabbit serum as diluents had no effect on the titer scores of the reaction, but using “normal” human AB serum as a diluent resulted in a lower titer score. Varying the ionic strength of the reacting system, we divided the sera into two groups. One group showed a greatly increased reactivity at 0.01 M, while the second showed severe prozone effects at that ionic strength. Using different pH levels at an ionic strength of 0.01 M showed that the cytotoxicity reactions at pH 6.5 and 7 were virtually identical but that the cytotoxicity reaction was undetectable at pH 8.