EpiQuik Total Histone H3 Quantification Kit (Fluorometric)

The EpiQuik™ Total Histone H3 Quantification Kit (Fluorometric) is a complete set of essential components which enables the experimenter to quantify the level of histone h3 protein independent of its modified state. The EpiQuik™ Total Histone H3 Quantification Kit can also be used for normalizing the modified histone H3 content of samples when run in parallel with Epigentek histone modification quantification kit series. 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 histone H3 with the detection limit as low as 2 ng/well and detection range from 10 ng-1 µg/well of histone extracts.
• The unmodified histone H3 control is conveniently included for the quantification of the amount of total histone H3.
• Strip microplate format makes the assay flexible: manual or high throughput.
• Simple, reliable, and consistent assay conditions.

Background Information
Histone H3- along with H2A, H2B, and H4- is involved in the structure of chromatin in eukaryotic cells. Histone H3 can undergo several different types of epigenetic modifications that influence cellular processes such as transcription activation/inactivation, chromosome packaging and DNA damage repair. These modifications including acetylation, phosphorylation, methylation, ubiquitination, and ADP-ribosylation, occur on the N-terminal tail domains of histone H3 through catalyzing of histone modifying enzymes, which results in remodeling of the nucleosome structure into an open conformation more accessible to transcription complexes. In most species, histone H3 is primarily acetylated at lysine 9, 14, 18, 23, and 56, and methylated at lysine 4, 9, 27, 36, and 79, and phosphorylated at ser10, ser28, Thr3, and Thr11, respectively. Thus, quantitative detection of various histone modifications would provide useful information for better understanding epigenetic regulation of cellular processes and for developing HMT-targeted drugs. Epigentek has provided a series of kits used for quantifying all sites/degrees of histone H3 modification. For added convenience and more quantitative interpretation of the results with use of these kits, we provide here the EpiQuik™ Total Histone H3 Quantification Kit (Fluorometric). This kit is designed for quantifying the level of histone H3 protein independent of its modified state and can also be used for normalizing the modified histone H3 content of samples when run in parallel with Epigentek histone modification quantification kit series.

Principle & Procedure
This kit is designed for quantifying the level of histone H3 protein independent of its modified state. In an assay with this kit, the histone H3 protein is captured to the strip-wells coated with an anti-histone H3 antibody. The captured histone H3 can then be detected with a detection antibody followed by a fluorescent development reagent. The ratio of histone H3 is proportional to the intensity of fluorescence. The absolute amount histone H3 can be quantified by comparing to the standard control.

Starting Materials
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.