The EpiQuik™ Global Pan-Methyl Histone H3-K79 Quantification Kit (Fluorometric) is a convenient package of tools that allows the experimenter to specifically measure global mono-, di-, and tri-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 3 hours.
- Innovative fluorometric assay without the need for radioactivity, electrophoresis, and chromatography.
- Simultaneous quantification of mono-, di-, and tri-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-, di-, and tri-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. H3-K79 di- and tri-methylation is also a widespread histone modification and is associated with transcriptionally active genes. Increased global H3-K79 methylation is also found to be involved in some pathological processes such as leukemogenesis in humans. The patterns of global H3-K79 methylation can also change by inhibition or activation of HMTs. Thus, quantitative detection of global mono-, di-, and tri-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 Pan-Methyl Histone H3-K79 Quantification Kit (Colorimetric) provides a tool for measuring mono-, di-, and tri-methylation of histone H3-K79.
Principle & Procedure
This kit is designed for simultaneously measuring global histone H3-K9 mono-, di-, and tri-methylation. In an assay with this kit, the methylated histone H3 at lysine 9 is captured to the strip wells coated with antibodies specifically for mono-, di-, and tri-methyl H3-K9. The captured mono-, di-, and tri-methylated histone H3-K9 can then be detected with a labeled detection antibody followed by a fluorescent development reagent. The ratio of mono-, di-, and tri-methylated H3-K9 is proportional to the intensity of fluorescence. The absolute amount of the methylated H3-K9 can be quantitated 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.