Revvity, 64PLKTPEG, HTRF Human Total PLK1 Detection Kit, 500 Assay Points

Overview
This HTRF cell-based assay enables the rapid, quantitative detection of total PLK1 protein (phosphorylated or not). Polo-like kinase 1 (PLK1) is the principle member of the well conserved serine/threonine kinase family and is essential for cell division (mitosis). As such, it is a key regulator of the G2/M checkpoint of the cell cycle.
PLK1 activation for cell cycle regulation: PLK1 is activated by phosphorylation on Thr210 by AurA kinase. Activated PLK1 phosphorylates CDC25.
PLK1 is a critical protein responsible for mitosis and cell cycle progression/cell proliferation. This makes it an attractive cancer treatment target.
How it works
Total PLK1 assay principle
The Total-PLK1 assay quantifies the expression level of PLK1 in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total-PLK1 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 PLK1 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 PLK1 two-plate assay protocol
The two-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 Total-PLK1 HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

 

Total PLK1 one-plate assay protocol
Detection of Total PLK1 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
Molecular Modification-Total
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target Class-Phosphoproteins
Target Species-Human
Technology-TR-FRET
Unit Size-500 assay points