The EpiQuik™ Global Mono-Methyl Histone H3-K79 Quantification Kit (Fluorometric) is a convenient package of tools that allows the experimenter to specifically measure global mono-methylation of histone H3-K79 fluorometrically using a variety of mammalian cells (human, mouse, etc.) including fresh and frozen tissues, cultured adherent and suspension cells. The kit has the following advantages:
- Quick and efficient procedure, which can be finished within 2.5 hours.
- Innovative fluorometric assay without the need for radioactivity, electrophoresis, and chromatography.
- Specifically captures mono-methylated H3-K79 with the detection limit as low as 0.4 ng/well and detection range from 5 ng-2 µg/well of histone extracts.
- The control is conveniently included for the quantification of the amount of mono-methylated H3-K79.
- Strip microplate format makes the assay flexible: manual or high throughput.
- Simple, reliable, and consistent assay conditions.
Epigenetic activation or inactivation of genes plays a critical role in many important human diseases, especially in cancer. A major mechanism for epigenetic inactivation of the genes is methylation of CpG islands in genome DNA caused by DNA methyltransferases. Histone methyltransferases (HMTs) control or regulate DNA methylation through chromatin-dependent transcription repression or activation. HMTs transfer 1-3 methyl groups from S-adenosyl-Lmethionine to the lysine and arginine residues of histone proteins. Dot1 is a histone methyltransferase that catalyzes methylation of histone H3 at lysine 79 (H3-K79) in mammalian cells. Intergenic H3-K79 mono-methylation is a mark of enhancer regions and may provide unique binding surfaces along the chromatin fiber to stabilize interactions between enhancers and their target promoters. The global H3-K79 mono-methylation can be changed by inhibiton or activation of HMTs. Thus quantitative detection of global mono-methyl histone H3-K79 would provide useful information for better understanding epigenetic regulation of gene activation and for developing HMT-targeted drugs. The EpiQuik™ Global Mono-Methyl Histone H3-K79 Quantification Kit (Fluorometric) provides a tool for measuring mono-methylation of histone H3-K79.
Principle & Procedure
This kit is designed for measuring global histone H3-K79 mono-methylation. In an assay with this kit, the mono-methylated histone H3 at lysine 79 is captured to the strip wells coated with an anti-monomethyl H3-K79 antibody. The captured mono-methylated histone H3-K79 can then be detected with a labeled detection antibody, followed by a fluorescent development reagent. The ratio of mono-methylated H3-K79 is proportional to the intensity of fluorescence. The absolute amount of mono-methylated H3-K79 can be quantified by comparing to the standard control.
Input material should be purified histone extracts. In general, the input amount should be from 50 ng to 200 ng per well of histone extracts.
F1 (10X Wash Buffer)
F2 (Antibody Buffer)
F3 (Detection Antibody, 1 mg/ml)*
F4 (Fluoro Developer)*
F5 (Fluoro Enhancer)*
F6 (Fluoro Dilution)
Standard Control (100 µg/ml)*
8-Well Sample Strips (with Frame)
8-Well Standard Control Strips
* For maximum recovery of the products, centrifuge the original vial prior to opening the cap.