EpiQuik Histone H3 Citrullination ELISA Kit (Colorimetric)

Product Description

The EpiQuik™ Histone H3 Citrullination ELISA Kit (Colorimetric) is a complete set of optimized buffers and reagents for specifically measuring global histone H3 citrullination from a broad range of species such as mammals, fungi, and bacteria, in a variety of forms including cultured cells and fresh tissues. Histone extracts can be prepared by using your own successful method. For your convenience and the best results, EpiGentek offers a histone extraction kit optimized for use with this kit. Histone extracts can be used immediately or stored at –80°C for future use. This kit has the following advantages:

• Quick and efficient procedure, which can be finished within 3.5 hours.
• High sensitivity and specificity. The detection limit is as low as 0.1 ng/well with dynamic range of  0.5-10 ng/well within the indicated amount range of the histone extracts. Only recognizes H3cit with no cross-reactivity with unmodified H3 or other modifications at the same arginine site.
• The control is conveniently included for the quantification of H3cit.
• Strip microplate format makes the assay flexible: manual or high throughput.
• Simple, reliable, and consistent assay conditions.

Background Information

Arginine histone citrullination is one of the many important epigenetic marks, and is essential for the regulation of epigenetic chromatin remodeling and immune cell’s extracellular trap processes. Citrullinated arginine of histone H3 (Arg2, 8, 17) regulates gene expression and is mainly catalyzed by peptidylarginine deiminase 4 (PAD4). Furthermore, citrullination of histones, in particular histone H3, was revealed as a convergence point for diverse inflammatory signals that trigger the neutrophil response to infections. It was reported that citrullinated histone H3 could be a potential serum biomarker for the early diagnosis of septic shock. More immunological diseases such as multiple sclerosis and rheumatoid arthritis seem to be also associated with change of histone H3 citrullination. The global histone H3 citrullination can be changed by inhibition or activation of PADs. Therefore, quantitative detection of global histone H3 citrullination would provide useful information for better understanding epigenetic regulation of gene activation and silencing, as well as for developing PAD-targeted drugs. 

Principle & Procedure

This kit is designed for measuring global H3cit. In an assay with this kit, the histone proteins are stably spotted on the strip wells. The citrulline H3 can be recognized with a high-affinity antibody and detected with a detection antibody, followed by a color development reagent. The ratio of H3cit is proportional to the intensity of absorbance. The absolute amount of H3cit can be quantitated by comparing to the standard control.

Starting Materials

Input materials should be histone extracts. The amount of histone extracts for each assay can be 50 ng to 200 ng with an optimal amount of 100 ng.

Performance Data

Fig.1. Histone extracts were prepared from HL-60 cells using the EpiQuik™ Total Histone Extraction Kit and the amount of H3cit was measured using the EpiQuik™ Histone H3 Citrullination ELISA Kit (Colorimetric).

Fig.2. Demonstration of high sensitivity and specificity of H3cit detection achieved by EpiQuik™ Histone H3 Citrullination ELISA Kit (Colorimetric). H3cit, unmodified H3 and methylated H3 at arginine site 2, 8 and 17, respectively were added into the assay wells at the different concentrations and then measured with the EpiQuik™ Histone H3 Citrullination ELISA Kit (Colorimetric).