Analysis Note

ControlIncluded negative control mouse IgG antibody and control primers specific for human RPL10 promoter.

Application

Research CategoryEpigenetics & Nuclear Function

Chromatin Immunoprecipitation:
Sonicated chromatin prepared from HeLa cells (1 X 10⁶ cell equivalents per IP) was subjected to chromatin immunoprecipitation using 2 µg of either a normal mouse IgG or Anti-acetyl-Histone H3 (Lys27) antibody and the Magna ChIP G (Cat. #17-611) Kit. Successful immunoprecipitation of acetyl-histone H3 (Lys27)-associated DNA fragments was verified by qPCR using β-globin Promoter ChIP Primers versus RPL10 Promoter Primers (Please see figures). 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 G ChIP™ (Cat. #17-408) or EZ-ChIP™ (Cat. #17-371) protocol for experimental details.

Western Blot Analysis:
Acid extracts from untreated (Lane 1) and sodium-butyrate treated (Lane 2) HeLa cells were resolved by electrophoresis, transferred to PVDF membrane and probed with Anti-acetyl Histone H3 (Lys27), clone CMA309 (0.1 µg/mL). Proteins were visualized using a goat
anti-mouse secondary antibody conjugated to HRP and a chemiluminescence detection system (Please see figures).

Research Sub CategoryChromatin Biology

Acetyl-Histone H3 (Lys27) ChIP validated antibody & primer set including the ChIP-grade antibody & the specific control PCR primers used for chromatin immunoprecipitation of H3K27Ac.

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

Lysine acetylation is a dynamic, reversible and tightly regulated protein and histone modification that plays a major role in chromatin remodeling and in the regulation of gene expression in various cellular functions. Histone acetylation is often associated with transcriptional activation and acetylation of H3K27 is found at active enhancers.

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.
The ChIPAb+ Acetyl-Histone H3 (Lys27) set includes the Anti-acetyl-Histone H3 (Lys27) antibody, a negative control antibody (purified Mouse IgG), and qPCR primers which amplify a 178 bp region within the promoter of the human RPL10 gene. The acetyl-histone H3 (Lys27) and negative control antibodies are supplied in a scalable "per ChIP" reaction size and can be used to functionally validate the precipitation of acetyl-histone H3 (Lys27)-associated chromatin.

Immunogen

The acetyl-histone H3 (Lys27) purified antibody is made against a synthetic peptide (acetylated at Lys27) corresponding to amino acids 19-37 of histone H3.

Epitope: a.a. 19-37

Legal Information

UPSTATE is a registered trademark of Merck KGaA, Darmstadt, Germany

Packaging

25 assays per kit, ~2µg per chromatin immunoprecipitation

Physical form

Format: Purified

Anti-acetyl-Histone H3 (Lys27) (mouse monoclonal IgG1, Clone CMA309). One vial containing 50 µg of protein G purified antibody in 50 µL PBS containing 0.05% sodium. Store at -20°C.

Normal Mouse IgG. Two vials containing 25 µg purified Mouse IgG in 25 µL storage buffer containing 0.1% sodium azide. Store at -20°C.

ChIP Primers, RPL10 Promoter. One vial containing 75 µL of 5 µM of each primer specific for the promoter region of human RPL10. Store at -20°C.
FOR: ACC CGT CTT CGA CAG GAC T
REV: GGA ACG GAA GAC GAG AAC AG

Quality

Chromatin Immunoprecipitation:
Sonicated chromatin prepared from HeLa cells (1 X 10⁶ cell equivalents per IP) were subjected to chromatin immunoprecipitation using 2 µg of either a normal mouse IgG or Anti-acetyl-Histone H3 (Lys27) antibody and the Magna ChIP G (Cat. #17-611) Kit.
Successful immunoprecipitation of acetyl-histone H3 (Lys27) associated DNA fragments was verified by qPCR using ChIP Primers RPL10 Promoter (Please see figures).
Please refer to the EZ-Magna G ChIP™ (Cat. #17-409) or EZ-ChIP™ (Cat. #17-371) protocol for experimental details.

Specificity

Recognizes histone H3, Mr 17 kDa, acetylated at Lys27.

This peptide sequence is identical in a wide range of animal and plant species.

Storage and Stability

Stable for 1 year at -20°C from date of receipt. Aliquot upon thawing, avoid freeze thaw cycles.

Target description

~17 kDa