Revvity, 64HDAC4TPEG, HTRF Human and Mouse Total HDAC4 Detection Kit, 500 Assay Points

Overview
Histone deacetylases (HDACs) regulate chromatin remodeling and subsequent gene transcription by controlling the status of histone acetylation. Histone deacetylation induces a condensed chromatin conformation, contributing to the repression of gene transcription which is involved in diverse physiological processes.
Importantly, HDACs are dysregulated in a number of brain disorders, which is implicated in the pathogenesis of these diseases, autism, Alzheimer's disease, and depressive disorders. It suggests that HDACs might be potential targets for the treatment of brain disorders.
Among class II HDACs, HDAC4 is a specific target for the treatment of ischemic stroke. It plays a key role in the pathogenesis of ischemic stroke and also in post-stroke recovery by affecting neuronal death, angiogenesis, and neurogenesis.
Epigenetic pathways define biologically relevant subsets of human cancers. EZH2 activation and HDAC4 activation correlate with growth factor signaling and inflammation, respectively, and represent two distinct states for cancer cells. This understanding may enable us to identify targetable drivers in breast cancer and mesenchymal glioblastoma.
How it works
Total HDAC4 assay principle
The Total-HDAC4 assay quantifies the expression level of HDAC4 in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total-HDAC4 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 HDAC4 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-HDAC4 2-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 Total HDAC4 HTRF detection reagents. This protocol enables the cells' viability and confluence to be monitored.

 

Total-HDAC4 1-plate assay protocol
Detection of total HDAC4 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 Mouse
Technology-TR-FRET
Unit Size-500 assay points