Tetramer technology: a technique in which MHC monomer molecules are tetramerized to increase their affinity and can be combined with multiple TCRs on T cells.
Introduction to tetramer technology
Since the soluble MHC monomer molecule has a low affinity with the TCR, the dissociation is fast, and the multivalent molecule can bind to multiple TCRs on one specific T cell, so that the dissociation speed is greatly slowed down. To this end, Altman et al. proposed the construction of MHC class I molecular tetramers by means of the biotin-avidin cascade amplification principle. The method uses a genetic engineering technique to add a 15 amino acid residue of Bio A substrate peptide (BSP) to a MHCI-like molecule such as the carboxy terminus of the HLA-A2 heavy chain to form a fusion protein, which is pressed in vitro. A certain proportion is incubated with beta microglobulin and specific antigenic short peptides to fold into the correct conformation and become pMHC complexes. Biotin is labeled on the lysine residue of the substrate peptide such that a fluorescein-labeled streptavidin binds to four biotinylated pMHC complexes to form a tetramer, MHC-antigen peptide tetramer and After binding of TCR on antigen-specific CFI-, antigen-specific CTLs in vivo can be quantified by flow cytometry and sorted for in vitro culture amplification and functional analysis.
The tetramer technology enables specific, efficient and direct quantification of antigen-specific CTI-activity assays, and can be applied to immunological research and detection, specific immunotherapy, and vaccine efficacy monitoring.
- As a clinical diagnostic tool, quantitatively measure the ratio of antigen-specific CTI in peripheral blood and tissues, and perform phenotypic and functional analysis. Altman et al. have detected antigen-specific CTI in a large number of asymptomatic AIDS patients. The detection rate is as high as 2%. Zerbini et al. used tetramer technology and immunohistochemistry to detect peptide-specific CD8 CTLs in tumor tissues of hepatocellular carcinoma patients for the first time, providing broad prospects for the application of immunotherapy to MAGE antigens in HCC. In situ staining of tetramers in autoimmune diseases was first used in TCR transgenic mice. Through the quantitative detection, phenotypic and functional analysis of antigen-specific CTL, it laid the foundation for elucidating the pathogenesis of viral infectious diseases, tumors and autoimmune diseases.
- For adoptive tumor immunotherapy: Mei-denbauer et al. administered a large number of Melan-A peptide-specific CTLs (42.1%) to 8 patients with refractory malignant melanoma, and found peripheral blood singles. The ratio of antigen-specific CTL in nuclear cells is determined by infusion. 01% to 0.07% rose to 2% after infusion. Moreover, these cells can survive in vivo for several weeks, have the function of secreting INF-7, and preferentially aggregate to the tumor to play a role.
(3) Monitoring of vaccine efficacy: In recent years, the use of antigen peptide pulse treatment as a vaccine to immunize the body to induce an effective CTL response has become a research hotspot. The efficacy of the vaccine can be monitored by tetramers constructed with homologous peptides.
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