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Comparison of peptide–major histocompatibility complex tetramers and dextramers for the identification of antigen-specific T cells
- Source :
- Clinical and Experimental Immunology
- Publication Year :
- 2014
- Publisher :
- Blackwell Science Inc, 2014.
-
Abstract
- Fluorochrome-conjugated peptide–major histocompatibility complex (pMHC) multimers are widely used for flow cytometric visualization of antigen-specific T cells. The most common multimers, streptavidin–biotin-based ‘tetramers’, can be manufactured readily in the laboratory. Unfortunately, there are large differences between the threshold of T cell receptor (TCR) affinity required to capture pMHC tetramers from solution and that which is required for T cell activation. This disparity means that tetramers sometimes fail to stain antigen-specific T cells within a sample, an issue that is particularly problematic when staining tumour-specific, autoimmune or MHC class II-restricted T cells, which often display TCRs of low affinity for pMHC. Here, we compared optimized staining with tetramers and dextramers (dextran-based multimers), with the latter carrying greater numbers of both pMHC and fluorochrome per molecule. Most notably, we find that: (i) dextramers stain more brightly than tetramers; (ii) dextramers outperform tetramers when TCR–pMHC affinity is low; (iii) dextramers outperform tetramers with pMHC class II reagents where there is an absence of co-receptor stabilization; and (iv) dextramer sensitivity is enhanced further by specific protein kinase inhibition. Dextramers are compatible with current state-of-the-art flow cytometry platforms and will probably find particular utility in the fields of autoimmunity and cancer immunology.
- Subjects :
- CD4-Positive T-Lymphocytes
Technology Advance
T cells
Biotin
Hemagglutinins, Viral
chemical and pharmacologic phenomena
T-Cell Antigen Receptor Specificity
Cell Separation
T cell receptors
CD8-Positive T-Lymphocytes
Cell Line
Major Histocompatibility Complex
HLA-A2 Antigen
Humans
Insulin
Protein Precursors
Telomerase
Fluorescent Dyes
diabetes
tumour immunology
autoimmunity
HLA-DR1 Antigen
Dextrans
Flow Cytometry
R1
Peptide Fragments
Clone Cells
Streptavidin
Protein Binding
Subjects
Details
- Language :
- English
- ISSN :
- 13652249 and 00099104
- Volume :
- 177
- Issue :
- 1
- Database :
- OpenAIRE
- Journal :
- Clinical and Experimental Immunology
- Accession number :
- edsair.pmid.dedup....60c99a29a0566474dcb32ff4d42ba5fc