Calorimetric studies of the state of water in deeply frozen human monocytes

Intra- and extracellular phase transitions in human peripheral blood monocyte suspensions with and without the cryoprotectant 1 M dimethylsulfoxide were measured using differential scanning calorimetry. Using an fluorescence diacetate/ethidium bromide assay for membrane integrity and a phagocytosis assay for cell function, it was found that mortality was correlated with several phase transitions under a variety of cooling and warming regimens. As a result of these studies we concluded that: intracellular freezing is lethal, but avoidance of freezing during fast cooling is not sufficient to provide complete protection; a subtle freezing injury in the cryoprotected monocytes can be correlated with a measurable increase in devitrification on warming; and the cell contents form more stable glasses than the Hanks' balanced salt solution with fetal calf serum used as the extracellular medium.