Journal
JOURNAL OF IMMUNOLOGICAL METHODS
Volume 268, Issue 1, Pages 51-69Publisher
ELSEVIER
DOI: 10.1016/S0022-1759(02)00200-4
Keywords
MHC tetramers; helper T cell; T lymphocyte; avidity; receptor-ligand interactions; multi-valent binding; flow cytometry; binding assay; HLA-DR
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Funding
- NIAID NIH HHS [AI 95361, K08 AI001698-01A1, AI 01698-01, AI 40873, K08 AI001698] Funding Source: Medline
- NIGMS NIH HHS [T32 GM08334] Funding Source: Medline
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Class I MHC-peptide oligomers (MHC tetramers) have become popular reagents for the detection and characterization of antigen-specific CD8(+) T cells. Class II MHC proteins can be produced by expression in Escherichia coli followed by in vitro folding, or by native expression in insect cells; biotin can be introduced by site-specific chemical modification of cysteine, or by enzymatic modification of a peptide tag; and a variety of fluorescent streptavidin preparations can be used for oligomerization. Here we review methodologies for production of fluorescent oligomers of soluble class II MHC proteins and discuss their use in analysis of antigen-specific CD4(+) T cells. We explore the experimental conditions necessary for efficient staining of CD4(+) T cells using oligomers of class II MHC proteins, and we establish a standard protocol. Finally, we consider complications and challenges associated with these reagents, discuss the interpretation of staining results, and suggest future. directions for investigation, in particular the use of MHC oligomers for the study of T cell avidity modulation. (C) 2002 Elsevier Science B.V. All rights reserved.
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