Revvity, 64METY34PEH, HTRF Human Phospho-MET (Thr1234/1235) Detection Kit, 10,000 Assay Points

Overview
c-MET, also called tyrosine-protein kinase MET or hepatocyte growth factor receptor (HGFR), is a human MET gene-coded protein with tyrosine kinase activity. It acts as a tyrosine kinase receptor, with functions related to wound healing, organogenesis, and embryonic development.
MET is expressed in epithelial cells and is triggered by hepatocyte factors HGF and HSF, with ramifications into various pathways like RAS-MAPK, AKT, and STAT3 signaling.
Abnormal activation of MET is linked to cancer progression, with effects on tumor growth, angiogenesis, and tumor cell migration & invasion. These relationships between MET and tumor cells are found across multiple cancer types (liver, brain, breast, kidney, etc) and are suspected to originate from a hijacking of MET and its ligand expression regulation, which leads to their overexpression and autocrine activation of MET.
How it works
HTRF Phospho MET Tyr1234/1235 assay principle
The HTRF Phospho MET Tyr1234/1235 assay measures MET when phosphorylated on the Tyrosine 1234/1235. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The assay uses 2 labeled antibodies: one with a donor fluorophore, the other with an acceptor. The first antibody was selected for its specific binding to the phosphorylated motif on the protein, and the second for its ability to recognize the protein independently of its phosphorylation state. Protein phosphorylation enables an immune-complex formation involving the two labeled antibodies which brings the donor fluorophore into close proximity to the acceptor, thereby generating a FRET signal. Its intensity is directly proportional to the concentration of phosphorylated protein present in the sample, and provides a means of assessing the protein’s phosphorylation state under a no-wash assay format.

 

HTRF Phospho MET Tyr1234/1235 2-plate assay protocol
The 2 plate protocol involves culturing cells in a 96-well plate before lysis, then transferring lysates into a 384-well low volume detection plate before the addition of the Phospho MET Tyr1234/1235 HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

 

HTRF Phospho MET Tyr1234/1235 one-plate assay protocol
Detection of phospho MET Tyr1234/1235 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 2
Molecular Modification-Phosphorylation
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target Class-Phosphoproteins
Target Species-Human
Technology-TR-FRET
Unit Size-10,000 assay points