Multiplexed labeling of viable cells for high-throughput analysis of glycine receptor function using flow cytometry

Gilbert, Daniel, Wilson, John C., Nink, Virginia, Lynch, Joseph W. and Osborne, Geoffrey W. (2009) Multiplexed labeling of viable cells for high-throughput analysis of glycine receptor function using flow cytometry. Cytometry Part A, 75A 5: 440-449. doi:10.1002/cyto.a.20703


Author Gilbert, Daniel
Wilson, John C.
Nink, Virginia
Lynch, Joseph W.
Osborne, Geoffrey W.
Title Multiplexed labeling of viable cells for high-throughput analysis of glycine receptor function using flow cytometry
Journal name Cytometry Part A   Check publisher's open access policy
ISSN 1552-4922
1552-4930
Publication date 2009-01-30
Year available 2009
Sub-type Article (original research)
DOI 10.1002/cyto.a.20703
Volume 75A
Issue 5
Start page 440
End page 449
Total pages 10
Editor Goolsby, Charles L.
Place of publication United States
Publisher John Wiley & Sons Inc
Collection year 2010
Language eng
Subject C1
110903 Central Nervous System
920111 Nervous System and Disorders
Abstract Flow cytometry is an important drug discovery tool because it permits high-content multiparameter analysis of individual cells. A new method dramatically enhanced screening throughput by multiplexing many discrete fixed cell populations; however, this method is not suited to assays requiring functional cellular responses. HEK293 cells were transfected with unique mutant glycine receptors. Mutant receptor expression was confirmed by coexpression of yellow fluorescent protein (YFP). Commercially available cell-permeant dyes were used to label each glycine receptor expressing mutant with a unique optical code. All encoded cell lines were combined in a single tube and analyzed on a flow cytometer simultaneously before and after the addition of glycine receptor agonist. We decoded multiplexed cells that expressed functionally distinct glycine receptor chloride channels and analyzed responses to glycine in terms of chloride-sensitive YFP expression. Here, data provided by flow cytometry can be used to discriminate between functional and nonfunctional mutations in the glycine receptor, a process accelerated by the use of multiplexing. Further, this data correlates to data generated using a microscopy-based technique. The present study demonstrates multiplexed labeling of live cells, to enable cell populations to be subject to further cell culture and experimentation, and compares the results with those obtained using live cell microscopy. © 2009 International Society for Advancement of Cytometry
Keyword multiplex
chloride channels
glycine receptor
Q-Index Code C1
Q-Index Status Confirmed Code

 
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