title: Loss of T cell responses following long-term cryopreservation.
abstract: Although cryopreservation of peripheral blood mononuclear cells (PBMC) is a commonly used technique, the degree to which it affects subsequent functional studies has not been well defined. Here we demonstrate that long-term cryopreservation has detrimental effects on T cell IFN-gamma responses in human immunodeficiency virus (HIV) infected individuals. Long-term cryopreservation caused marked decreases in CD4(+) T cell responses to whole proteins (HIV p55 and cytomegalovirus (CMV) lysate) and HIV peptides, and more limited decreases in CD8(+) T cell responses to whole proteins. These losses were more apparent in cells stored for greater than one year compared to less than six months. CD8(+) T cell responses to peptides and peptide pools were well preserved. Loss of both CD4(+) and CD8(+) T cell responses to CMV peptide pools were minimal in HIV-negative individuals. Addition of exogenous antigen presenting cells (APC) did not restore CD4(+) T cell responses to peptide stimulation and partially restored T cell IFN-gamma responses to p55 protein. Overnight resting of thawed cells did not restore T cell IFN-gamma responses to peptide or whole protein stimulation. A selective loss of phenotypically defined effector cells did not explain the decrement of responses, although cryopreservation did increase CD4(+) T cell apoptosis, possibly contributing to the loss of responses. These data suggest that the impact of cryopreservation should be carefully considered in future vaccine and pathogenesis studies. In HIV-infected individuals short-term cryopreservation may be acceptable for measuring CD4(+) and CD8(+) T cell responses. Long-term cryopreservation, however, may lead to the loss of CD4(+) T cell responses and mild skewing of T cell phenotypic marker expression.
authors: Blood Systems Research Institute, 270 Masonic Avenue, San Francisco, CA 94118, USA. rowen@bloodsystems.org
authors: Rachel E
authors: RE
authors: Owen
authors: Elizabeth
authors: E
authors: Sinclair
authors: Brinda
authors: B
authors: Emu
authors: John W
authors: JW
authors: Heitman
authors: Dale F
authors: DF
authors: Hirschkorn
authors: C Lorrie
authors: CL
authors: Epling
authors: Qi Xuan
authors: QX
authors: Tan
authors: Brian
authors: B
authors: Custer
authors: Jeffery M
authors: JM
authors: Harris
authors: Mark A
authors: MA
authors: Jacobson
authors: Joseph M
authors: JM
authors: McCune
authors: Jeffery N
authors: JN
authors: Martin
authors: Frederick M
authors: FM
authors: Hecht
authors: Steven G
authors: SG
authors: Deeks
authors: Philip J
authors: PJ
authors: Norris
mesh_terms: D000208:Acute Disease; D017209:Apoptosis / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology; D002460:Cell Line; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D002908:Chronic Disease; D018920:Coculture Techniques; D015925:Cryopreservation; D003587:Cytomegalovirus / Q000276:immunology; D003586:Cytomegalovirus Infections / Q000276:immunology / Q000473:pathology; D006678:HIV / Q000276:immunology; D015658:HIV Infections / Q000276:immunology / Q000473:pathology; D006801:Humans; D008297:Male; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D013997:Time Factors
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