Revvity, 64NACPEH, HTRF Human and Mouse Total ACC Detection Kit, 10,000 Assay Points

Overview
ACC (Acetyl-CoA carboxylase) is an important regulator of fatty acid metabolism. By catalyzing the carboxylation of acetyl-CoA to malonyl-CoA, ACC serves as a novel biomarker or drug target in diabetes and obesity research. The enzymatic activity of ACC is inhibited by phosphorylation on Ser-79 residue.
Like various genes involved in fatty acid regulation, the transcription of ACC genes is tightly regulated by the SREBP1c transcription factor. When activated, this transcription factor binds to the SRE DNA sequence upstream from the ACC gene promoter, and activates its transcription. Thus, the quantitative detection of Total ACC represents a valuable readout of SREBP1c pathway activation.
The HTRF Total ACC cellular assay monitors the expression levels of both ACC 1 and ACC2, and can be used as a normalization assay with the HTRF phospho-ACC Ser79 kit. This kit is compatible with the buffers from the phospho-ACC kit, so the same lysate can be used for analyzing both the phosphorylated and the total protein populations.
This cell-based assay enables simple yet sensitive and efficient quantification of ACC 1/2, and offers enhanced convenience over ELISA or WB assays.
How it works
Total-ACC assay principle
The Total-ACC assay quantifies the expression level of ACC in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total-ACC 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 the presence of ACC 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-ACC 2-plate assay protocol
The 2 plate protocol involves culturing cells in a 96-well plate before lysis, then transferring lysates to a 384-well low volume detection plate before the addition of Total ACC HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

 

Total-ACC 1-plate assay protocol
Detection of total ACC 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 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
Therapeutic Area-Metabolism/Diabetes NASH/Fibrosis
Unit Size-10,000 assay points