Single‐Cell Phospho‐Protein Analysis by Flow Cytometry
Open Access
- 15 August 2007
- journal article
- unit
- Published by Wiley in Current Protocols in Immunology
- Vol. 78 (1), 8.17.1-8.17.20
- https://doi.org/10.1002/0471142735.im0817s78
Abstract
This protocol describes methods for monitoring intracellular phosphorylation-dependent signaling events on a single-cell basis. This approach measures cell signaling by treating cells with exogenous stimuli, fixing cells with formaldehyde, permeabilizing with methanol, and then staining with phospho-specific antibodies. Thus, cell signaling states can be determined as a measure of how cells interact with their environment. This method has applications in clinical research as well as mechanistic studies of basic biology. In clinical research, diagnostic or drug efficacy information can be retrieved by discovering how a disease affects the ability of cells to respond to growth factors. Basic scientists can use this technique to analyze signaling events in cell lines and human or murine primary cells, including rare populations, like B1 cells or stem cells. This technique has broad applications to take standard biochemical analysis into primary cells to garner valuable information about signaling events in physiologic settings. Curr. Protoc. Immunol. 78:8.17.1-8.17.20. © 2007 by John Wiley & Sons, Inc.Keywords
This publication has 28 references indexed in Scilit:
- The ERK Mitogen-Activated Protein Kinase Pathway Contributes to Ebola Virus Glycoprotein-Induced CytotoxicityJournal of Virology, 2007
- K-RasG12D expression induces hyperproliferation and aberrant signaling in primary hematopoietic stem/progenitor cellsBlood, 2006
- Flt3 Y591 duplication and Bcl-2 overexpression are detected in acute myeloid leukemia cells with high levels of phosphorylated wild-type p53Blood, 2006
- Altered B-cell receptor signaling kinetics distinguish human follicular lymphoma B cells from tumor-infiltrating nonmalignant B cellsBlood, 2006
- Fluorescent cell barcoding in flow cytometry allows high-throughput drug screening and signaling profilingNature Methods, 2006
- Transcending the biomarker mindset: deciphering disease mechanisms at the single cell levelCurrent Opinion in Chemical Biology, 2006
- T-Cell Tropism and the Role of ORF66 Protein in Pathogenesis of Varicella-Zoster Virus InfectionJournal of Virology, 2005
- Whole blood fixation and permeabilization protocol with red blood cell lysis for flow cytometry of intracellular phosphorylated epitopes in leukocyte subpopulationsCytometry Part A, 2005
- Detection of Intracellular Phosphorylated STAT-4 by Flow CytometryClinical Immunology, 2001
- Detection of Intracellular Phosphorylated STAT-1 by Flow CytometryClinical Immunology, 1999