Product Pathways - Cytoskeletal Signaling
Alexa Fluor® 647 Phalloidin #8940
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Alexa Fluor® 647 Phalloidin allows researchers to fluorescently stain the cytoskeleton through the binding of phalloidin to F-actin. This product is intended for use on fixed and permeabilized samples due to the toxicity associated with phalloidin. After reconstitution the stock solution provides enough material to perform 300 assays based on a 1:20 dilution and a 100 μl assay volume.
Alexa Fluor® 647 Fluorescent Properties: Excitation: 650, Emission: 668.
Actin, a ubiquitous eukaryotic protein, is the major component of the cytoskeleton. At least six isoforms are known in mammals. Nonmuscle β- and γ-actin, also known as cytoplasmic actin, are predominantly expressed in nonmuscle cells, controlling cell structure and motility (1). Actin exists mainly as a fibrous polymer, F-actin. In response to cytoskeletal reorganizing signals during processes such as cytokinesis, endocytosis, or stress, cofilin promotes fragmentation and depolymerization of F-actin, resulting in an increase in the monomeric globular form, G-actin (2). Phalloidin is a natually occuring toxic bicyclic peptide found in the deathcap toadstool, Amanita phalloides, that rapidly binds to F-actin with strong affinity (3).
Directions For Use
To make a stock concentration of 6.6 μM (20X), reconstitute the lyophilized material in 1.5 ml pure methanol.
Fix cells for 15 minutes using fresh, methanol-free 4% formaldehyde, then rinse three times in PBS for 5 minutes each. Once fixed, please follow CST protocol for immunostaining. Following incubation of the primary and secondary antibodies, Alexa Fluor® 647 Phalloidin can be diluted 1:20 (5 μl per 100 μl assay volume) in PBS and added to cells. Allow to incubate for 15 minutes at room temperature, then rinse once with PBS. Coverslip slides with ProLong® Gold antifade reagent and examine specimen using appropriate excitation wavelength.
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For Research Use Only. Not For Use In Diagnostic Procedures.
Alexa Fluor® is a registered trademark of Molecular Probes, Inc.
Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.