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STAR FAK ELISA Kit
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PRODUCT FAMILY INFORMATION
STAR ELISA Kits
Upstate STAR (Signal Transduction Assay Reaction) ELISA kits are a fast, sensitive method to detect the relative amounts of total and activated targets through the use of phospho-specific antibodies.
Upstate STAR (Signal Transduction Assay Reaction) ELISA kits are a fast, sensitive method to detect the relative amounts of total and activated targets through the use of phospho-specific antibodies.
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Description:
STAR FAK ELISA Kit
Trade Name:
Upstate (Millipore)
Qty/Pk:
96 assays
Background Information:
I. TEST PRINCIPLE
The UPSTATE® colorimetric STAR (Signal Transduction Assay Reaction) ELISA kit is a solid phase sandwich enzyme linked immunosorbent assay that provides a fast, sensitive method to detect specific levels of signaling targets in whole cell extracts. The FAK plate is coated with a specific mouse monoclonal FAK capture antibody. Sample lysate or the standard included in the kit are incubated in the microwells allowing FAK antigen to be captured. The plate is then washed to remove any un-bound non-specific material. A specific rabbit anti-FAK antibody is added to detect the captured FAK on the plate. The unbound detection antibody is washed away followed by incubation with an HRP-conjugated anti-rabbit antibody. After the addition of TMB substrate and stop solution the absorbance is measured at 450 nm using a plate reader. This allows for a sensitive enzymatic detection of the sample.
The entire assay takes less than 5 hours to complete with minimal hands-on time. Many of the reagents are supplied in ready-to use formulations for ease of use. The kit also includes a standard that is run as both a positive control and to generate a standard curve for FAK measurement.
II. FAK BACKGROUND
FAK (Focal Adhesion Kinase) is a non-receptor tyrosine kinase which localizes to focal adhesions and plays a key role in integrin-mediated signaling pathways regulating cell migration. It autophosphorylates on tyrosine 397 during adhesion and spreading, and is thought to be a critical transducer of adhesion-dependent growth and survival. FAK and it phosphorylation states have been implicated in cancer metastasis and tumor cell survival and adhesion-independent growth. Additionally, recent evidence indicates that elevation of FAK activity in human carcinoma cells is associated with increased invasive potential. A central role in tumor formation and progression suggest that FAK is an attractive target for therapeutic intervention.
Cell surface integrins when engaged with their ligands lead to the recruitment of a number of intracellular proteins to specialized sites of the cytoplasmic face into focal adhesions. These focal adhesion plaques are composed of many proteins that include kinases, phosphatases, scaffolding proteins, and G-proteins. The activation of integrins leads to the tyrosine phosphorylation of several cytoplasmic proteins critical to the biochemical process, including FAK, Src, paxillin, tensin, and p130Cas. This clustering and activation of proteins at the membrane leads to the activation of many diverging signaling pathways that result in cytoskeletal re-organization. This starts with either the recruitment of Src and FAK to the plasma membrane following integrin or PI3 Kinase activation. The non-receptor protein-tyrosine kinases (PTKs) FAK and Src are key members of the focal adhesion complex. These PTKs co- Localize and associate with the transmembrane integrins and various downstream targets of certain growth factor receptors. The outcome of Src binding to FAK is the phosphorylation of several tyrosine residues in FAK and its related proteins, including the FAK autophosphorylation site Tyr397. This autophosphorylation occurs immediately after integrin clustering and allows for the associat |