Detection, sorting, and immortalization of dual expresser lymphocytes from human peripheral blood samples

STAR Protoc. 2021 Oct 30;2(4):100925. doi: 10.1016/j.xpro.2021.100925. eCollection 2021 Dec 17.

Abstract

This protocol describes how to identify Dual Expressers (DEs), a rare type of lymphocyte that co-expresses B-cell receptors and T-cell receptors, by flow cytometry using a cocktail of four antibodies. It also shows the subsequent gating strategy for detecting and sorting DEs and the generation of EBV-immortalized DE lymphoblastoid cell lines and clones for antibody production and cloning antigen receptors. Use of this protocol maximizes detection of DEs and minimizes inclusion of doublets. For complete details on the use and execution of this protocol, please refer to Ahmed et al. (2019).

Keywords: Antibody; Cell Biology; Cell culture; Cell isolation; Flow Cytometry/Mass Cytometry; Immunology.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adolescent
  • Adult
  • Aged
  • Aged, 80 and over
  • Blood Cells / cytology
  • Cell Culture Techniques / methods*
  • Cell Line
  • Centrifugation, Density Gradient
  • Child
  • Female
  • Flow Cytometry / methods*
  • Humans
  • Immunohistochemistry
  • Lymphocytes* / chemistry
  • Lymphocytes* / classification
  • Lymphocytes* / cytology
  • Lymphocytes* / metabolism
  • Male
  • Middle Aged
  • Receptors, Antigen, B-Cell / chemistry
  • Receptors, Antigen, B-Cell / metabolism
  • Receptors, Antigen, T-Cell / chemistry
  • Receptors, Antigen, T-Cell / metabolism
  • Young Adult

Substances

  • Receptors, Antigen, B-Cell
  • Receptors, Antigen, T-Cell