Analysis of Inflammasome Activation in Response to Yersinia Infection by Fluorescence Microscopy Detection of Active Caspase-1 Puncta.

Analysis of Inflammasome Activation in Response to Yersinia Infection by Fluorescence Microscopy Detection of Active Caspase-1 Puncta. Methods Mol Biol. 2019;2010:231-240 Authors: Zwack EE, Brodsky IE Abstract The type of cell death triggered by a particular environmental stimulus influences the outcome of infection or inflammatory disease processes. The ability to identify the cell death pathway that is activated in response to infection is essential for understanding the pathogenesis and host response to infection. Activation of the cysteine protease caspase-1 in various inflammasome complexes indicates that cells are undergoing pyroptosis, a regulated, proinflammatory cell death. Inflammasome assembly and caspase activation can be measured by various methods ranging from detection of inflammasome-dependent cell death, cytokine secretion, cleavage of caspase-1, or the formation of "puncta" within the cell that contain inflammasome components, such as caspase-1 or the adapter protein ASC. Here we describe a method for detecting caspase-1 activation on a single cell level in the context of infection by the Gram-negative pathogen Yersinia using immunofluorescence microscopy. We previously used this approach to quantify caspase-1 puncta formation in cells containing Yersinia translocon components (Zwack et al., MBio 6:e02095-14, 2015). This is a modification of methods used previously by Broz et al. (Cell Host Microbe 8:471-483, 2010) ...
Source: Mol Biol Cell - Category: Molecular Biology Authors: Tags: Methods Mol Biol Source Type: research