ChIPAb+ Phospho-Histone H3 (Ser28)

Stock Code: 3586379
Manufacturer Part No: 17-10269

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Descriptions

Analysis Note

ControlIncludes normal rabbit IgG and primers specific for human GAPDH promoter

Application

Research Sub CategoryHistones

Chromatin Immunoprecipitation:
Representative lot data.
Sonicated chromatin prepared from untreated or colcemid-treated HeLa cells (1 X 106 cell equivalents per IP) were subjected to chromatin immunoprecipitation using either 5 µl Normal rabbit IgG, or 5 µl Anti-phospho-Histone H3 (Ser28) and the Magna ChIP^® A Kit (Cat. # 17-610). Successful immunoprecipitation of phospho-Histone H3 (Ser28)- associated DNA fragments was verified by qPCR using ChIP Primers, human GAPDH promoter region for untreated and treated chromatin samples (Figure 2). Data is presented as percent input of each IP sample relative to input chromatin for each amplicon and ChIP sample as indicated.
Please refer to the EZ-Magna ChIP™ A (Cat. # 17-408) or EZ-ChIP™ (Cat. # 17-371) protocol for experimental details.
Western Blot Analysis:
Representative lot data.
HeLa acid extract (lane 1) and recombinant Histone H3 (lane 2) were probed with Anti-phospho-Histone H3 (Ser28) (1:2,000 dilution).
Proteins were visualized using a Donkey Anti-Rabbit IgG secondary antibody conjugated to HRP and a chemiluminescence detection system. (Figure 3)

Arrow indicates phospho-Histone H3 (Ser28) (~17 kDa).
Dot Blot Specificity Analysis:
Representative lot data.
Histone peptides with various modifications (see table) were probed with Anti-phospho-Histone H3 (Ser28). Proteins were visualized using a Donkey Anti-Rabbit IgG secondary antibody conjugated to HRP and a chemiluminescence detection system. (Figure 4)
Immunocytochemistry Analysis
Representative lot data
Confocal fluorescent analysis of HeLa cells using Anti-phospho-Histone H3 (Ser28) (Red). Actin filaments have been labeled with AlexaFluor™ 488 dye-Phalloidin (Green). Nucleus is stained with DAPI (Blue). This antibody positively stains the mitotic cell nucleus. (Figure 5)
Peptide Inhibition Assay: A 1:500 dilution from a representative lot was blocked by phospho-Histone H3 (Ser28) peptides, but not non-specific peptides.

Research CategoryEpigenetics & Nuclear Function

This ChIPAb+ Phospho-Histone H3 (Ser28) -ChIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.

Disclaimer

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

General description

All ChIPAb+ antibodies are individually validated for chromatin precipitation, every lot, every time. Each ChIPAb+ antibody set includes control primers (tested every lot by qPCR) to biologically validate your IP results in a locus-specific context. The qPCR protocol and primer sequences are provided, allowing researchers to validate ChIP protocols when using our antibody in their chromatin context. Each set also includes a negative control antibody to ensure specificity of the ChIP reaction.