Revvity, 64CDK9TPEG, HTRF Human and Mouse Total CDK9 Detection Kit, 500 Assay Points

Overview
The Total CDK9 cellular assay monitors CDK9 protein levels.
CDK9 in association with cyclins T or K, also called the positive transcription elongation factor (P-TEFb), is a general transcription factor that regulates transcription elongation through phosphorylation of the C-terminal tail domain (CTD) of RNA polymerase II (RNAP II). This crucial function and the evidence that transcriptional programs are dysregulated in cancer made make CDK9 an attractive target for anti-cancer therapies.
HTRF assays offer many advantages over other technologies:

Homogeneous add-and-read format
No wash steps
Low background
Straightforward miniaturization from 96- or 384-well microplates to high density assay formats such as 384-well low volume and 1536-well plates
Stable signal, providing flexibility in time of readout or size of assays
How it works
Total CDK9 assay principle
The Total CDK9 assay quantifies the expression level of CDK9 in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total CDK9 assay uses two labeled antibodies, one coupled to a donor fluorophore and the other to an acceptor. Both antibodies are highly specific for a distinct epitope on the protein. In the presence of CDK9 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 CDK9 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 Total CDK9 HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

 

Total CDK9 one-plate assay protocol
Detection of Total CDK9 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
Automation Compatible-Yes
Brand-HTRF
Detection Modality-HTRF
Lysis Buffer Compatibility-Lysis Buffer 1 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 Mouse
Technology-TR-FRET
Therapeutic Area-Oncology & Inflammation
Unit Size-500 assay points