title: Effects of cryopreservation on effector cells for antibody dependent cell-mediated cytotoxicity (ADCC) and natural killer (NK) cell activity in (51)Cr-release and CD107a assays.
abstract: Freshly isolated PBMC are broadly used as effector cells in functional assays that evaluate antibody-dependent cell mediated cytotoxicity (ADCC) and NK activity; however, they introduce natural-individual donor-to-donor variability. Cryopreserved PBMC provide a more consistent source of effectors than fresh cells in cytotoxicity assays. Our objective was to determine the effects of cryopreservation of effector PBMC on cell frequency, and on the magnitude and specificity of ADCC and NK activity. Fresh, frozen/overnight rested and frozen/not rested PBMC were used as effector cells in (51)Cr-release and CD107a degranulation assays. Frozen/overnight rested PBMC had higher ADCC and NK activity in both assays when compared to fresh PBMC; however, when using frozen/not rested PBMC, ADCC and NK activities were significantly lower than fresh PBMC. Background CD107a degranulation in the absence of target cell stimulation was greater in PBMC that were frozen/not rested when compared to fresh PBMC or PBMC that were frozen overnight and rested. The percentages of CD16(+)CD56(dim) NK cells and CD14(+) monocytes were lower in PBMC that were frozen and rested overnight than in fresh PBMC. CD16 expression on CD56(dim) NK cells was similar for all PBMC treatments. PBMC that were frozen and rested overnight were comparable to fresh PBMC effectors. PBMC that were frozen and used immediately when evaluating ADCC or NK activity using either a (51)Cr-release assay or a CD107a degranulation assay had the lowest activity. Clinical studies of antibodies that mediate ADCC would benefit from using effector cells that have been frozen, thawed and rested overnight prior to assay.
authors.affiliation: Department of Immunology/Microbiology, Rush University Medical Center, Chicago, IL 60612, United States.
authors.forename: Mariana M
authors.initials: MM
authors.lastname: Mata
authors.affiliation: Department of Immunology/Microbiology, Rush University Medical Center, Chicago, IL 60612, United States.
authors.forename: Fareeha
authors.initials: F
authors.lastname: Mahmood
authors.affiliation: Department of Immunology/Microbiology, Rush University Medical Center, Chicago, IL 60612, United States.
authors.forename: Ryan T
authors.initials: RT
authors.lastname: Sowell
authors.affiliation: Department of Immunology/Microbiology, Rush University Medical Center, Chicago, IL 60612, United States. Electronic address: Linda_L_Baum@rush.edu.
authors.forename: Linda L
authors.initials: LL
authors.lastname: Baum
mesh_terms: D000920:Antibody-Dependent Cell Cytotoxicity / Q000276:immunology*; D016175:B-Lymphocyte Subsets / Q000276:immunology*; D019002:CD56 Antigen / Q000378:metabolism; D015550:Cell Degranulation / Q000276:immunology; D002460:Cell Line; D002860:Chromium Radioisotopes / Q000032:analysis; D015925:Cryopreservation; D003601:Cytotoxicity Tests, Immunologic / Q000379:methods*; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology*; D018950:Lipopolysaccharide Receptors / Q000378:metabolism; D052118:Lysosomal-Associated Membrane Protein 1 / Q000032:analysis; D017452:Receptors, IgG / Q000378:metabolism
chemical_list: D019002:CD56 Antigen; D002860:Chromium Radioisotopes; D018950:Lipopolysaccharide Receptors; D052118:Lysosomal-Associated Membrane Protein 1; D017452:Receptors, IgG
keywords: ADCC; CD107a; Cryopreservation; Degranulation; NK cells; PBMC