Revvity, 64RNAP2TPEG, HTRF Human and Mouse Total RNA Polymerase II Detection Kit, 500 Assay Points

Overview
RNA polymerase II (RNA Pol II) is one of the primary enzymes responsible for expressing protein-encoding genes and some small nuclear RNAs. The carboxy-terminal domain (CTD) of RNA Pol II undergoes a cycle of phosphorylation, catalyzed mainly by members of the cyclin-dependent kinase (CDK) family. RNA Pol II CTD phosphorylation allows it to temporally couple transcription with transcription-associated processes (binding specificity of the CTD for particular factors is determined by the site-specific phosphorylation/dephosphorylation events).
How it works
Total RNA Pol II assay principle
The Total RNA Pol II assay quantifies the expression level of RNA Pol II in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total RNA Pol II 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 RNA Pol II 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 RNA Pol II 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 HTRF Total RNA Pol II detection reagents. This protocol enables the cells' viability and confluence to be monitored.

Total RNA Pol II one-plate assay protocol
Detection of Total RNA Pol II 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-Protein Quantification
Brand-HTRF
Buffer/Solvent-Lysis Buffer 1
Detection Modality-HTRF
Host Species-Human Mouse
Molecular Modification-Total
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target-RNA pol II
Target Class-Phosphoproteins
Target Species-Human Mouse
Technology-TR-FRET
Therapeutic Area-Immuno-oncology Inflammation
Unit Size-500 assay points