PMID: 9433460Jan 20, 1998Paper

Simultaneous detection of DNA synthesis and cytokine production in staphylococcal enterotoxin B activated CD4+ T lymphocytes by flow cytometry

Journal of Immunological Methods
B A Mehta, V C Maino

Abstract

Assessment of T cell activation has traditionally been performed by measuring proliferation as a function of 3[H]-thymidine incorporation, or secretion of cytokines from activated peripheral blood mononuclear cells (PBMC) in culture. An alternative method for detection of proliferation at the single cell level utilizes incorporation of bromodeoxyuridine (BrdU), an analog of thymidine, into cellular DNA. After appropriate fixation and permeabilization of the cells, a monoclonal antibody (mAb) against BrdU conjugated with a fluorescent dye is employed to measure by flow cytometry the incorporated BrdU. Here, we report a flow cytometric procedure which can be used for the simultaneous detection of BrdU incorporation, activation markers such as CD69 and CD25, and intracellular cytokines in T cell subsets from activated PBMC. Our observations are consistent with the proposal that cytokine synthesis and cell proliferation occur sequentially in CD4+ T cells stimulated with the superantigen staphylococcal enterotoxin B (SEB). The majority of cells expressing the cytokines IFN-gamma and IL-2 at 48 h appear to have undergone DNA synthesis, however all proliferating cells do not express IFN-gamma or IL-2. The methods presented in this rep...Continue Reading

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Citations

May 13, 2004·Clinical Immunology : the Official Journal of the Clinical Immunology Society·Andreas ThielAndreas Radbruch
Jan 17, 2002·Transplantation·Benny J ChenNelson J Chao
Apr 2, 2003·Thorax·V BrownM Ennis
Nov 30, 2000·Arthritis Research·A Thiel, A Radbruch
Feb 13, 2001·Journal of Neuroimmunology·S MuhallabA Svenningsson
Aug 19, 2004·The Journal of Allergy and Clinical Immunology·Thomas A Fleisher, João Bosco Oliveira
May 20, 2004·International Journal of Medical Microbiology : IJMM·Matthew D Baker, K Ravi Acharya
Aug 31, 2010·Journal of Immunological Methods·Andreia SoaresWillem A Hanekom
Dec 12, 2012·Critical Reviews in Microbiology·Ram Sarup Singh, Amandeep K Walia
Oct 25, 2002·Cytotherapy·E D Wieder
May 17, 2002·The Journal of Veterinary Medical Science·Maki MotobuYoshikazu Hirota
May 9, 2000·Memórias do Instituto Oswaldo Cruz·M A SantiagoS G Coutinho
Jun 10, 2005·International Immunology·Yi JiangJonathan H Williams
Dec 23, 2003·The Journal of Immunology : Official Journal of the American Association of Immunologists·Matthew J Loza, Bice Perussia

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