Slow versus Fast Postthaw Dilution for the Recovery of Lymphocytes in Whole Blood Frozen in either Dimethyl Sulfoxide or Glycerol

Abstract

It was previously shown by the authors that human whole blood was cryoprotected by dimethyl sulfoxide (DMSO) for use as inoculum in short-term lymphocyte cultures. That procedure utilized a fast postthaw dilution where the cryoprotective agent was diluted in two steps. In this paper it is shown that a five-step ‘slow’ dilution technique is much superior to the two-step ‘fast’ dilution procedure, which significantly revises our original report of 1972. This method has strikingly improved the degree of blastogenic response to phytohemagglutinin as measured by ³H-thymidine incorporation. The increase in amount of tritium incorporation was seen in cultures which were protected with either DMSO or glycerol. A similar effect was observed in unfrozen control cultures exposed to either cryoprotectant.