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For the Ex Vivo Quantification of Cellular Immune Response
T lymphocytes play a central role in immune system function. Total T cell and T cell subset counts are measured by detection of various cell surface molecules. Refined enumeration of CD8+ antigen-specific T cells requires cognate recognition of the T cell receptor by a Class I MHC (Major Histocompatibility Complex) / peptide complex. This can be done using Class I MHC Tetramers.1,2
MHC Tetramers are complexes of four MHC (Major Histocompatibility Complex) molecules, which are associated with a specific peptide and bound to a fluorochrome. These complexes bind to a distinct set of T cell receptors (TCRs) on a subset of CD8+ T cells.2 Thus, mixing tetramers with PBLs or whole blood and detection using flow cytometry, a count of all T cells that are specific for one peptide and its matched allele is provided, regardless of functionality. Simply stated, iTAg reagents will allow us to measure the cellular response against one specific peptide. These tetramers, which have been mutated to minimize binding of HLA to cell surface CD8, show a greatly diminished binding to the general population of CD8 cells, but retain peptide-specific binding, thus facilitating accurate discrimination of rare, specific T cells (less than 1% of CD8+).3,4
"...the new generation of HLA-tetramers bind the CTLs with a sensitivity down to 1 in 5,000 CD8+ T cells."
-- G.S. Ogg and A.J. McMichael.Current Opinion in Immunology. 1998.4
Allele and Peptide Specificity
The iTAgTM MHC Tetramers recognize human CD8+ T cells which are specific for the peptide and HLA molecule in the complex. The complex is composed of four HLA-A*0201 MHC molecules, each bound to the specific peptide and conjugated with phycoerythrin (PE). The HLA-A*0201 allele is found in 40% - 50% of the global population. The HLA molecule in the reagents has been modified to minimize CD8 mediated binding.3
Specific binding does not depend on functional pathways. Therefore, the population identified by these tetramers includes all specific CD8+ cells regardless of functional status. With all available reagents, total CD8 events may be enumerated by staining with the CD8-FITC antibody.
Proven through extensive research that has been documented in over 100 publications, MHC Tetramer technology has been used to accurately and efficiently monitor CD8+ T cell responses in the following conditions: Viruses (Human Immunodeficiency Virus-Acquired Immunodeficiency Syndrome, Epstein Barr virus-mononucleosis, cytomegalovirus, human papilloma virus, hepatitis B, hepatitis C, influenza and measles) Parasitic infection (malaria) Cancer (breast, prostate, melanoma, colon, lung, cervical) Autoimmune diseases (multiple sclerosis, rheumatoid arthritis) Transplantation
Identifying the Most Appropriate iTAg MHC Tetramers
New, ready-to-use flow cytometry reagents include Class I, HLA-A* 0201, MHC Tetramers / CD8 HIV pol, HIV gag, Mart1, EBV BMLF1, Influenza M1, and CMV pp65. Please click here for more information or click below to order products.
Custom Tetramer Solutions are suitable for basic and clinical research in a number of applications including drug / vaccine development. Beckman Coulter will manufacture Class I A*0201 tetramers incorporating peptides specified by the customer for specific research uses. Click here to learn more about how Custom iTAg MHC Tetramers can support your research.
To review the benefits of iTAg MHC Tetramer technology, click here. For more information on iTAg MHC Tetramers, visit the Frequently Asked Questions section.
"Recent technological advances have shed unexpected light on the CD8+ T cell responses to pathogens and have raised concerns that a familiar quantitative assay is wrong. The most exciting of the new advances is the MHC tetramer."
-- A.J. McMichael and C.A. O'Callaghan. Journal of Experimental Medicine. 1998.2
References:
- Altman JD, Moss PH, Goulder PJ, et al. Phenotypic analysis of antigen-specific T lymphocytes. Science. 1996; 274: 94-96.
- McMichael AJ, O'Callaghan CA. A new look at T cells. J Exp Med. 1998; 187: 1367-1371.
- Bodinier M, Peyrat M-A, Tournay C, et al. Efficient detection and immunomagnetic sorting of specific T cells using multimers of MHC class I and peptide with reduced CD8 binding. Nat Med. 2000; 6: 707-710.
- Ogg GS, McMichael AJ. HLA-peptide tetrameric complexes. Curr Opin Immunol. 1998; 10: 393-396.
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