Combinatorial color coding

When using MHC multimers to study antigen-specific T cells, each MHC/peptide detects only a small fraction of T cells that are specific for that peptide.  Thus, for a large screening of several specificities a proportionally large volume of blood or sample will be needed. To address this issue, an excellent strategy is to use a "combinatorial color coding"1. The method consist in assigning a unique two-color code to each tetramer/peptide of interest. Two tetramers tagged with different colors will bind to the TCRs, yielding a specific T cell that can be detected in two channels in a flow cytometer. By analyzing the double positive cells it is possible to identify the peptide that those cells are recognizing.

The number of tetramer/peptide combinations that can be used simultaneously depend on how many fluorophores are used to assemble the tetramers. Using 5 different streptavidin-fluorophore conjugates allows for 10 unique two-color combinations (light blue region, table 1). Using 6 conjugates, the codes expand to 15 (light red region, table 1). The codes increase as the number of conjugates increase. Thus, as you increase the colors used, more specificities can be covered and less sample is required. The use of BioLegend's Flex-Tâ„¢ technology allows for an efficient and affordable solution for antigen-specific T cell multi screening. The cost of the reagents is much lower than other commercially available products. In addition, we offer in-house, custom screening. Please contact our Custom Solution Team for a quote: cst@biolegend.com.

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Table 1. Possible combinations of two-color codes depend on the number of Streptavidin-fluorophore conjugates used to generate the tetramer/peptide complex

Table 1. Possible combinations of two-color codes depend on the number of Streptavidin-fluorophore conjugates used to generate the tetramer/peptide complex

1) Hadrup SR et al. Parallel detection of antigen-specific T-cell responses by multidimensional encoding of MHC multimers. Nat Methods. 2009 Jul;6(7):520-6.