BioTechniques (Mar 2017)

Preparation of peptide–MHC and T-cell receptor dextramers by biotinylated dextran doping

  • Michael T. Bethune,
  • Begoña Comin-Anduix,
  • Yu-Hsien Hwang Fu,
  • Antoni Ribas,
  • David Baltimore

DOI
https://doi.org/10.2144/000114525
Journal volume & issue
Vol. 62, no. 3
pp. 123 – 130

Abstract

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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.

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