Revvity, 64COFTPEH, HTRF Human and Mouse Total Cofilin Detection Kit, 10,000 Assay Points

Overview
The kit is designed for the rapid detection of Total Cofilin in cell supernatant and whole cells. Actin-binding proteins are abundant cellular proteins that regulate cell function by mediating actin polymerization and remodeling. Cofilin, known as a regulator of actin filament dynamics, is a small ~21kDA protein that is ubiquitously expressed in all vertebrates and freely diffuses in eukaryotic cells.
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 Cofilin assay principle
The HTRF Total Cofilin assay quantifyies the expression level of Cofilin. It is entirely plate-based and does not require gels, electrophoresis, or transfer. The assay uses two antibodies, one coupled to a donor fluorophore and the other to an acceptor. The two antibodies are highly specific for a distinct epitope on the protein. In presence of Cofilin 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 Cofilin two-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 Total Cofilin HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

 

Total Cofilin one-plate assay protocol
Detection of Total Cofilin 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 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
Unit Size-10,000 assay points