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17-479 STAR FAK ELISA Kit

This STAR FAK ELISA Kit is used to measure & quantify FAK levels.

STAR FAK ELISA Kit MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.
Key Applications: ELISA Detection Methods: Chromogenic Research Category: Epigenetics & Nuclear Function, Cell Signaling
17-479
96 assays  
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      Overview

      Replacement Information

      Key Spec Table

      Analytes AvailableSpecies ReactivityKey ApplicationsDetection Methods
      FAK H, M, RELISAChromogenic
      Description
      Catalogue Number17-479
      Brand Family Upstate
      Trade Name
      • STAR
      • Upstate
      DescriptionSTAR FAK ELISA Kit
      Alternate Names
      • Focal adhesion kinase 1
      • FADK 1
      • Focal adhesion kinase-related nonkinase
      • FRNK
      • Protein phosphatase 1 regulatory subunit 71
      • PPP1R71
      • Protein-tyrosine kinase 2
      • p125FAK
      • pp125FAK
      Background InformationI. 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 association of FAK with various signaling proteins such as PI3 kinase, Grb2-Sos, p130Cas, and paxillin. The phosphorylation of FAK Tyr397 is crucial for many of the established biological roles of FAK, including cell migration, cell cycle progression, and prevention of anoikis, a form of detachment-induced apoptosis and is correlated with the invasiveness of tumors and is thought to be a pivotal player by modulating the turnover of focal adhesions to regulate cell migration. Phosphoinositide 3-Kinase (PI3 kinase or PI3K) also plays a role in cell migration via its binding to the autophosphorylation site (Tyr397) of FAK through one or both of its SH2 domains in the p85 subunit of PI3K. This is the same site in FAK that is bound by Src. In theory, the binding of PI3 kinase to FAK may be involved in activities like cell proliferation, apoptosis, and migration that are related to the phosphorylation of Tyr397. This is substantiated by the observation that AKT is downstream of PI3 kinase and is a mediator in preventing apoptosis. Therefore it is possible that FAK/PI3 kinase association may help regulate apoptosis.
      Materials Required but Not Delivered1. Multi-channel or repeating pipettes
      2. Plate shaker (optional)
      3. Pipettors and tips capable of accurately measuring 1-1000 µL
      4. Graduated serological pipettes
      5. 96-well microtiter Plate Reader with 450 nm filter
      6. Graphing software for plotting data or graph paper for manual plotting of data
      7. Microfuge tubes for standard and sample dilutions
      8. Mechanical vortex
      9. 1 liter container
      10. Distilled or deionized water
      References
      Product Information
      Components
      • Capture Plate pre-coated with anti-FAK antibody: (Part No. 17-479A) One pre-coated 96-stripwell immunoplate sealed in a foil pouch.
      • Anti-FAK detection antibody: (Part No. 17-479B) One bottle (11 mL) of anti-FAK detection antibody containing sodium azide, ready to use.
      • ELISA Diluent: (Part No. 17-479C) One bottle (25 mL) of ELISA Diluent containing sodium azide, ready to use.
      • 25X ELISA Wash Buffer: (Part No. 17-479D) One bottle (50 mL) of 25X ELISA Wash Buffer.
      • Anti-Rabbit IgG HRP conjugate: (Part No. 17-479E) One vial (125 µL) of 100X anti-rabbit HRP conjugate containing thimerosal.
      • HRP Diluent: (Part No. 17-479F) One bottle (25 mL) of HRP Diluent containing thimerosal.
      • TMB Solution: (Part No. 17-479G) One bottle (25 mL) of stabilized tetramethylbenzidine (TMB), ready to use.
      • Stop Solution: (Part No. 17-479H) One bottle (25 mL) of stop solution, ready to use.
      • FAK Standard: (Part No. 17-479I) Two vials of FAK standard, lyophilized.
      • Plate Covers: Two plate covers.
      Detection methodChromogenic
      Applications
      ApplicationThis STAR FAK ELISA Kit is used to measure & quantify FAK levels.
      Key Applications
      • ELISA
      Biological Information
      Species Reactivity
      • Human
      • Mouse
      • Rat
      Analytes Available
      • FAK
      Entrez Gene Number
      Entrez Gene SummaryThis gene encodes a cytoplasmic protein tyrosine kinase which is found concentrated in the focal adhesions that form between cells growing in the presence of extracellular matrix constituents. The encoded protein is a member of the FAK subfamily of protein tyrosine kinases but lacks significant sequence similarity to kinases from other subfamilies. Activation of this gene may be an important early step in cell growth and intracellular signal transduction pathways triggered in response to certain neural peptides or to cell interactions with the extracellular matrix. At least four transcript variants encoding four different isoforms have been found for this gene, but the full-length natures of only two of them have been determined.
      Gene Symbol
      • PTK2
      • pp125FAK
      • FADK
      • FAK1
      • FAK
      UniProt Number
      UniProt SummaryFUNCTION: SwissProt: Q05397 # Non-receptor protein-tyrosine kinase implicated in signaling pathways involved in cell motility, proliferation and apoptosis. Activated by tyrosine-phosphorylation in response to either integrin clustering induced by cell adhesion or antibody cross-linking, or via G-protein coupled receptor (GPCR) occupancy by ligands such as bombesin or lysophosphatidic acid, or via LDL receptor occupancy. Plays a potential role in oncogenic transformations resulting in increased kinase activity.
      SIZE: 1052 amino acids; 119233 Da
      SUBUNIT: Interacts with CAS family members and with GIT1, SORBS1 and BCAR3. Interacts with SHB (By similarity). Interacts with TGFB1I1.
      SUBCELLULAR LOCATION: Cell junction, focal adhesion; Cytoplasmic side. Note=Constituent of focal adhesions.
      TISSUE SPECIFICITY: Expressed in all organs tested, in lymphoid cell lines, but most abundantly in brain.
      DOMAIN: SwissProt: Q05397 The first Pro-rich domain interacts with the SH3 domain of CRK-associated substrate (BCAR1) and CASL (By similarity). & The carboxy-terminal region is the site of focal adhesion targeting (FAT) sequence which mediates the localization of FAK1 to focal adhesions.
      PTM: Phosphorylated on 6 tyrosine residues upon activation.
      SIMILARITY: Belongs to the protein kinase superfamily. Tyr protein kinase family. FAK subfamily. & Contains 1 FERM domain. & Contains 1 protein kinase domain.
      Physicochemical Information
      Sensitivity
      • Sensitivity: 1.6 ng/mL
        Range of Detection: 1.6 to 100 ng/mL
        Species Reactivity: Human, mouse and rat
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Usage Statement
      • Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
      Storage and Shipping Information
      Storage ConditionsMaintain the unopened kit at 2-8°C until expiration date.

      Precautions

      • The instructions provided have been designed to optimize the kit's performance. Deviation from the instructions may result in suboptimal performance of the kit and the failure to produce accurate data.
      • Caustic Material: Stop Solution. Caution: Eye, hand, face, and clothing protection should be worn when handling this material.
      • Safety Warnings and Precautions: This kit is designed for research use only and not recommended for internal use in humans or animals. All chemicals should be considered potentially hazardous and principles of good laboratory practice should be followed.
      • The Detection Antibody and ELISA Diluent contain sodium azide. Sodium azide may react with copper and lead plumbing to form highly explosive metal azides. Upon disposal, flush with large amounts of water to prevent azide build-up. Avoid contact with skin.
      • The Anti-Rabbit IgG HRP Conjugate and HRP Diluent contain thimerosal. Thimerosal is highly toxic by inhalation, contact with skin or if swallowed. Thimerosal is a possible mutagen and should be handled accordingly.
      Packaging Information
      Material Size96 assays
      Transport Information
      Supplemental Information
      Specifications
      Global Trade ITEM Number
      Catalogue Number GTIN
      17-479 04053252741111

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      Categories

      Life Science Research > Antibodies and Assays > Immunoassays > Enzyme-linked Immunosorbent Assay (ELISA) > Complete ELISA Kits
      Life Science Research > Protein Detection and Quantification > Immunoassays > Enzyme-linked Immunosorbent Assay (ELISA) > Complete ELISA Kits