Enrichment & Staining

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Cellular and subcellular visualization is often hampered by the presence of background signal and staining. This is especially problematic when interrogating the dynamics of a low abundance protein. Often times, an enrichment step is required to reduce background and amplify the desired signal.

MilliporeSigma’s enrichment and staining kits not only stain highly abundant structural features of the cell, but also enrich for desired proteins and organelles bound to the actin cytoskeleton.

ProteoExtract® Cytoskeleton Enrichment and Staining Kit
Immunofluorescence Analysis: Confocal fluorescence microscopy of non-treated/enriched and treated/enriched HeLa cells. (A) F-actin was detected using TRITC-conjugated, (B) focal adhesion contacts were detected using a Vinculin antibody and a FITC-conjugated secondary, (C) nuclear counterstaining revealed with DAPI and all images were overlaid. Background due to soluble cytosolic fraction is significantly reduced after enrichment using the ProteoExtract Cytoskeleton Enrichment and Staining Kit (Cat. No. 17-10210), resulting in clear detection of insoluble, low-abundance actin-associated proteins (vinculin).
The actin cytoskeleton serves as a scaffold to assemble multi-component signaling complexes. Upon activation, many actin regulatory proteins/phospho-proteins move from the soluble cytoplasmic compartment to the insoluble actin cytoskeleton. The insolubility of these important proteins makes it difficult to study their biochemical changes upon binding to actin including regulatory post-translational modifications (phosphorylation, nitrosylation, etc.) 

MilliporeSigma’s easy-to-use cytoskeleton enrichment kit quickly and gently enriches the actin cytoskeleton while maintaining its native, adhered conformation, with minimal disruption of pre-existing protein associations. The kit allows for the visualization and co-localization of the actin cytoskeleton and actin-associated proteins in their native state and reduces the background typically associated with traditional methods of whole cell staining, greatly increasing the ability to detect low abundance actin-associated proteins.

For biochemical analysis, see our ProteoExtract Cytoskeleton Enrichment and Isolation Kit (Catalogue No. 17-10195).

Actin Cytoskeleton / Focal Adhesion Staining Kit
The organization of actin filaments and focal adhesions reflects cell polarity, cell cycle state, differentiation status, and other phenotypes.
Immunocytochemistry Analysis: Confocal fluorescence microscopy of focal adhesion and actin cytoskeleton in COS-7 cells revealed with triple labeling using TRITC-conjugated phalloidin, anti-Vinculin and DAPI.
The organization of actin filaments and focal adhesions reflects cell polarity, cell cycle state, differentiation status, and other phenotypes. This staining kit is a sensitive immunocytochemical tool to map the local orientation of actin filaments and focal adhesions relative to the nucleus. This kit consists of three components (TRITC-conjugated phalloidin, anti-Vinculin and DAPI) for the immunofluorescent staining of actin filaments in the cytoskeleton, focal contacts as well as the nucleus of the cells. Reagents and materials supplied in this kit are sufficient for 100 tests (including controls).