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J Immunol. 2015 Jan 1;194(1):455-62. doi: 10.4049/jimmunol.1401110. Epub 2014 Nov 17.

A novel flow cytometric method to assess inflammasome formation.

Author information

1
School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Queensland 4072, Australia; katryn.stacey@uq.edu.au d.sester@uq.edu.au.
2
School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Queensland 4072, Australia;
3
Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland 4072, Australia; and.
4
Queensland Brain Institute, University of Queensland, Brisbane, Queensland 4072, Australia.
5
School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Queensland 4072, Australia; Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland 4072, Australia; and katryn.stacey@uq.edu.au d.sester@uq.edu.au.

Abstract

Inflammasomes are large protein complexes induced by a wide range of microbial, stress, and environmental stimuli that function to induce cell death and inflammatory cytokine processing. Formation of an inflammasome involves dramatic relocalization of the inflammasome adapter protein apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) into a single speck. We have developed a flow cytometric assay for inflammasome formation, time of flight inflammasome evaluation, which detects the change in ASC distribution within the cell. The transit of ASC into the speck is detected by a decreased width or increased height of the pulse of emitted fluorescence. This assay can be used to quantify native inflammasome formation in subsets of mixed cell populations ex vivo. It can also provide a rapid and sensitive technique for investigating molecular interactions in inflammasome formation, by comparison of wild-type and mutant proteins in inflammasome reconstitution experiments.

PMID:
25404358
DOI:
10.4049/jimmunol.1401110
[Indexed for MEDLINE]
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