Cryopreservation of human granulocytes: study of granulocyte function and ultrastructure

Human granulocytes can be cryopreserved with dimethyl sulfoxide (DMSO) at--80 degrees C. However, the percent recovery of functional cells has been unsatisfactory to date. Throughout this study we have been able to prepare relatively pure granulocytes approximately equal to 85%), cryopreserve them with 5%--10% DMSO with and without serum, and store them at--80 degrees C for up to 4 mo. The parameters studied were absolute cell counts and viability determination, myeloperoxidase activity, phagocytosis, candidacidal activity, bactericidal activity, nitroblue tetrazolium reduction, chemiluminescence, and cell morphology by transmission and scanning electron microscopy. Based on our investigation, granulocytes cryopreserved without serum showed an intact membrane of superior integrity as compared with those preserved with serum. At least 50% of the cells recovered were functional after 2 mo of storage, but there was a progressive loss of viability and function on prolonged storage. The property of phagocytosis was the best preserved after storage for 4 mo, whereas myeloperoxidase activity, killing activity, nitroblue tetrazolium reduction, and chemiluminescence were maintained less efficiently. Morphological studies of cryopreserved granulocytes revealed that the nuclear, cytoplasmic, and cell surface architectures were altered by storage. Depletion of nuclear and cytoplasmic material, as well as changes in configuration, were also noted.