Preparation of peptide–MHC and T-cell receptor dextramers by biotinylated dextran doping
| Autor: | Michael T. Bethune, Begoña Comin-Anduix, Yu-Hsien Hwang Fu, Antoni Ribas, David Baltimore |
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| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: | |
| Zdroj: | BioTechniques, Vol 62, Iss 3, Pp 123-130 (2017) |
| Druh dokumentu: | article |
| ISSN: | 00011452 1940-9818 0736-6205 |
| DOI: | 10.2144/000114525 |
| Popis: | Peptide–major histocompatibility complex (pMHC) multimers enable the detection, characterization, and isolation of antigen-specific T-cell subsets at the single-cell level via flow cytometry and fluorescence microscopy. These labeling reagents exploit a multivalent scaffold to increase the avidity of individually weak T-cell receptor (TCR)-pMHC interactions. Dextramers are an improvement over the original streptavidin-based tetramer technology because they are more multivalent, improving sensitivity for rare, low-avidity T cells, including self/tumor-reactive clones. However, commercial pMHC dextramers are expensive, and in-house production is very involved for a typical biology research laboratory. Here, we present a simple, inexpensive protocol for preparing pMHC dextramers by doping in biotinylated dextran during conventional tetramer preparation. We use these pMHC dextramers to identify patient-derived, tumor-reactive T cells. We apply the same dextran doping technique to prepare TCR dextramers and use these novel reagents to yield new insight into MHC I–mediated antigen presentation. |
| Databáze: | Directory of Open Access Journals |
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