Histone H3 phosphorylation Info
Histone modifications such as phosphorylation, acetylation and methylation at specific amino acid residues on the histone tails that extend beyond the core nucleosome have been found to influence and regulate transcription, chromosome packaging and DNA damage repair. Phosphorylation of serine 10 and serine 28 in the tail of histone H3 occur early in mitosis when chromosomes begin to condense and during premature chromosome condensation induced in S-phase cells. Histone H3 is phosphorylated on serine 10 (pS10) during late S phase or G2 phase, while the phosphorylation of serine 28 (pS28) occurs during prophase. In contrast to serine 10, serine 28 phosphorylation has never been observed in interphase.
Histone H3 phospho Ser28 ELISA (H3S28ph)
The Histone H3 phospho Ser28 ELISA is capable of detecting the level of histone H3 phosphorylated at serine 28 from core histone preparations or histones purified by acid extraction from tissue or cell samples (Figure 1).
Figure 1: Histone H3 phospho Ser28 ELISA (H3 pSer28).
The Histone H3 phospho Ser28 ELISA was used to assay 78 ng to 5 µg of HeLa acid extracts that were either untreated (Catalog No. 36200) or paclitaxel treated (Catalog No. 36201). Data shown are the results from wells assayed in duplicate. These results are provided for demonstration only.
The Histone Modification ELISA Advantage
Historically, screening for histone modifications from sample preparations have been conducted using immunoblotting and chromatin immunoprecipitation methods, which can be time-consuming, do not allow for high-throughput and provide only semi-quantitative results. Active Motif's Histone Modification ELISAs provide a fast, sensitive assay with the flexibility to screen from 1 to 96 samples in a single experiment.
Included in each Histone Methylation ELISA is a methylated recombinant histone protein made using Active Motif's patented protein synthesis technology. This protein can be used to build a reference standard curve to quantitate the amount of specifically methylated histone H3 in your samples. The Total Histone H3 ELISA includes an unmodified Recombinant Histone H3 protein. Each Histone Phosphorylation ELISA contains treated and untreated acid extracts for use as a positive and negative controls.
Why use the Histone Methylation and Histone Phosphorylation ELISAs?
- Increased sensitivity over immunoblotting methods
- Results in less than three hours
- Specific antibody detection ensures low background and no cross-reactivity with other modifications
- Colorimetric readout enables easy, quantitative analysis with spectrophotometry at 450 nm
- 96-stripwell format enables both high and low throughput
- Positive controls included in each kit
The Histone Methylation ELISA Method
The Histone ELISAs to detect site- and degree-specific lysine methylation levels on histone H3 are sandwich ELISAs that utilize a monoclonal histone H3 antibody to capture histone H3 from purified core histones or histones isolated by acid extraction from tissue or cell samples. A polyclonal antibody specific for the modification of interest is used for detection, while a secondary antibody conjugated to horseradish peroxidase (HRP) and developing solutions provide a sensitive colorimetric readout that is easily quantified by spectrophotometry at 450 nm. The assay is performed in a convenient 96-stripwell plate, which enables either low or high throughput screening.
The Histone Phosphorylation ELISA Method
The Histone ELISAs to detect phosphorylation levels of serine 10 (pSer10) or serine 28 (pSer28) on histone H3 are sandwich ELISAs that utilize a C-terminal histone H3 antibody to capture histone H3 from purified core histones or histones isolated by acid extraction from tissue or cell samples. A monoclonal antibody specific for the phosphorylation site of interest is used for detection, while a secondary antibody conjugated to horseradish peroxidase (HRP) and developing solutions provide a sensitive colorimetric readou