Revvity, 64NAKPEG, HTRF Human and Mouse Total FAK Detection Kit, 500 Assay Points

Overview
The Total FAK cellular assay monitors total FAK and is used as a normalization assay with the phospho-FAK kit. This kit is compatible with the buffers from the phospho-FAK kit, so the same lysate can be used for analyses of both the phosphorylated and the total protein populations.
FAK, Focal Adhesion Kinase, is a cytoplasmic tyrosine kinase controlling cell adhesion, migration, proliferation, and survival. FAK is a central regulator of ECM/integrin and cadherin signaling, and plays a role in cell adhesion, cell junction, migration, survival, and mechanosensing. FAK can be activated by Growth Factor Receptors, Cytokine Receptors, and GPCRs. Increased FAK expression and phosphorylation is associated with increased tumor cell adhesion, migration, invasion in several cancers, and with fibroblast migration on fibronectin in tissue fibrosis.
How it works
Total-FAK assay principle
The Total FAK assay quantifies the expression level of FAK in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total-FAK assay uses two labeled antibodies, one coupled to a donor fluorophore, the other to an acceptor. Both antibodies are highly specific for a distinct epitope on the protein. In presence of FAK in a cell extract, the addition of these conjugates brings the donor fluorophore into close proximity with the acceptor and thereby generates a FRET signal. Its intensity is directly proportional to the concentration of the protein present in the sample, and provides a means of assessing the protein's expression under a no-wash assay format.

Total-FAK two-plate assay protocol
The two-plate protocol involves culturing cells in a 96-well plate before lysis, then transferring lysates to a 384-well low volume detection plate before the addition of Total-FAK HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

Total-FAK one-plate assay protocol
Detection of Total FAK with HTRF reagents can be performed in a single plate used for culturing, stimulation, and lysis. No washing steps are required. This HTS designed protocol enables miniaturization while maintaining robust HTRF quality. Application-Cell Signaling
Brand-HTRF
Detection Modality-HTRF
Lysis Buffer Compatibility-Lysis Buffer 1 Lysis Buffer 2 Lysis Buffer 3 Lysis Buffer 4 Lysis Buffer 5
Molecular Modification-Total
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target Class-Phosphoproteins
Target Species-Human Mouse
Technology-TR-FRET
Therapeutic Area-Cardiovascular NASH/Fibrosis Oncology & Inflammation
Unit Size-500 assay points