@inbook{70a32f74a3974eb9be6d2608274d58c2,
title = "Measuring NLR oligomerization II: Detection of ASC speck formation by confocal microscopy and immunofluorescence",
abstract = "Inflammasome assembly results in the formation of a large intracellular protein scaffold driven by the oligomerization of the adaptor protein apoptosis-associated speck-like protein containing a CARD (ASC). Following inflammasome activation, ASC polymerizes to form a large singular structure termed the ASC “speck,” which is crucial for recruitment of caspase-1 and its inflammatory activity. Hence, due to the considerably large size of these structures, ASC specks can be easily visualized by microscopy as a simple upstream readout for inflammasome activation. Here, we provide two detailed protocols for imaging ASC specks: by (1) live-cell imaging of monocyte/macrophage cell lines expressing a fluorescently tagged version of ASC and (2) immunofluorescence of endogenous ASC in cell lines and human immune cells. In addition, we outline a protocol for increasing the specificity of ASC antibodies for use in immunofluorescence.",
keywords = "ASC, Confocal microscopy, Flow cytometry, Immunofluorescence, Inflammasome, Live-cell imaging, Speck",
author = "Michael Beilharz and {De Nardo{\textquoteright}S}, Dominic and Eicke Latz and Franklin, {Bernardo S.}",
year = "2016",
month = jan,
day = "1",
doi = "10.1007/978-1-4939-3566-6_9",
language = "English",
isbn = "9781493935642",
series = "Methods in Molecular Biology",
publisher = "Humana Press",
pages = "145--158",
editor = "{Di Virgilio}, {Francesco } and Pablo Pelegrin",
booktitle = "NLR Proteins",
address = "United States of America",
}