부산대학교 도서관

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.jim.2007.02.003 Byline: Marta Bull (a), Deborah Lee (b), Jason Stucky (b), Ya-Lin Chiu (c), Abbe Rubin (c), Helen Horton (b)(d), M. Juliana McElrath (b)(d)(e) Keywords: T cells; IFN-gamma; Cryopreservation Abstract: Interferon-gamma (IFN-[gamma]) ELISpot and intracellular cytokine staining (ICS) assays are routinely employed in clinical HIV vaccine trials to identify antigen-specific T cells in cryopreserved peripheral blood mononuclear cells (PBMC). Several parameters involved in blood collection, processing and shipping may influence immunological function of the resulting cells, including anticoagulant type, time from venipuncture to PBMC isolation/cryopreservation, method of PBMC isolation and procedure for sample shipping. We examined these parameters in single and multiple site studies, and found the length of time from venipuncture to cryopreservation is the most important parameter affecting performance of T cells in immunological assays. Comparing blood processed at 24 h after venipuncture with that processed within 8 h, we observed on average a modest reduction in PBMC viability ([approximately equal to]8% decrease), a greater loss in cell recovery ([approximately equal to]32%), and between 36-56% loss in IFN-[gamma] T cell frequencies by ELISpot assay. We also describe three cold shipping methods that maintain immunological function in appropriately cryopreserved PBMC. These data indicate that cryopreservation of PBMC should occur within 8 h of venipuncture for optimal performance. This narrow window for specimen processing has important implications in selecting and monitoring clinical sites with laboratory capacity to perform these procedures in future clinical trials. Author Affiliation: (a) Children's Hospital and Regional Medical Center, Seattle, WA, United States (b) Program in Infectious Diseases, Fred Hutchinson Cancer Research Center (FHCRC), Seattle, WA, United States (c) Statistical Center for HIV/AIDS Research and Prevention (SCHARP), Fred Hutchinson Cancer Research Center (FHCRC), Seattle, WA, United States (d) Department of Medicine, University of Washington School of Medicine, Seattle, WA, United States (e) Laboratory Medicine, University of Washington School of Medicine, Seattle, WA, United States Article History: Received 16 August 2006; Revised 5 January 2007; Accepted 1 February 2007