A rapid microassay for mixed lymphocyte interactions (MLI) in the mouse and the guinea pig has been developed. This assay is based on the stimulation of protein synthesis (SPS) that occurs as soon as 11 hr after initiation of the culture and can be measured with high precision. The cells are cultured in leucine-free medium without serum in microtiter plates in 0.2 ml volumes using 2 × 106 cells of each cell type. Stimulation of protein synthesis as measured by increased incorporation of a pulse of 3H-leucine into protein occurs in allogeneic MLI's in which cells differ at major and minor, as well as at only minor histocompatibility loci, but does not occur in xenogeneic MLI's. Unidirectional MLI's can also be performed using either Xirradiated cells or cells from F1-hybrid animals. The sum of the two one-way MLI's in each case equals that of the two-way MLI. The advantage of this assay are: (1) it can be completed in less than 24 hr; (2) serum is not required in the culture medium; (3) relatively few cells and small amounts of reagents are required; (4) it is highly reproducible.
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