title: Characterization of purified cryopreserved human monocyte function in assays of superoxide production, accessory cell function, chemotaxis, and in fluorescent cell sorter analysis.
abstract: The ability to use highly purified cryopreserved human monocytes in various in vitro assays has a number of practical and theoretical advantages, including convenience and the potential for enhanced reproducibility. Large numbers (up to 1 X 10(9)) human monocytes can be isolated from a single donor in a purified suspension state, by a combination of leukapheresis and counter-current centrifugal elutriation (CCE) technologies. Following short- and long-term periods of cryopreservation, the viability and phagocytic function of these CCE-purified monocytes was unimpaired. Cryopreserved monocytes were similar to fresh cells in their ability to release superoxide anion (O2-), although unstimulated and stimulated O2- release values tended to increase slightly following weeks to months of cryopreservation. In contrast, even short-term cryopreservation diminished both the random migration and chemotactic responses of human monocytes; however, cryopreserved monocytes could be employed in this assay provided the calculated chemotactic ratio (chemotactic migration/spontaneous migration) was used. Cryopreserved monocytes demonstrate 70% of fresh monocyte accessory cell function in pokeweed mitogen-induced lymphocyte proliferation assays. When the binding of OKT3, OKM1, anti-DR, and fluoresceinated pokeweed mitogen to monocytes was analyzed in the fluorescence activator cell sorter (FACS), cryopreserved and fresh monocytes displayed a similar pattern of membrane reactivity.
mesh_terms: D002634:Chemotaxis, Leukocyte; D005434:Flow Cytometry / Q000379:methods; D005615:Freezing; D006801:Humans; D007937:Leukapheresis; D008213:Lymphocyte Activation; D008214:Lymphocytes / Q000276:immunology; D009000:Monocytes / Q000166:cytology* / Q000276:immunology / Q000502:physiology; D011309:Preservation, Biological; D013481:Superoxides / Q000097:blood*
chemical_list: D013481:Superoxides