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ChIPAb+ Trimethyl-Histone H3 (Lys4) - ChIP Validated Antibody and Primer Set
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REFERENCES
The organization of histone H3 modifications as revealed by a panel of specific monoclonal antibodies.
Kimura, Hiroshi, et al. (2008) Cell Struct. Funct., 33: 61-73 (2008)
Species Reactivity Key Applications Host Format Antibody Type
H, Vrt WB, ICC, ChIP Mouse Protein G Purified Monoclonal Antibody
Description:
ChIPAb+ Trimethyl-Histone H3 (Lys4) - ChIP Validated Antibody and Primer Set
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Trade Name:
Upstate (Millipore)
Product Overview:
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+ Trimethyl-Histone H3 (Lys4) set includes the Anti-trimethyl-Histone H3 (Lys4) antibody, a negative control antibody (purified Mouse IgG), and qPCR primers which amplify a 166 bp region within the promoter of the human GAPDH gene. The trimethyl-Histone H3 (Lys4) and negative control antibodies are supplied in a scalable "per ChIP" reaction size and can be used to functionally validate the precipitation of trimethyl-histone H3 (Lys4)-associated chromatin.
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Specificity:
Recognizes histone H3, Mr 17 kDa, trimethylated at Lys4.
Molecular Weight:
Trimethyl histone H3 at ~17 kDa
Epitope:
a.a. 1-12
Immunogen:
The trimethyl-histone H3 (Lys4) purified antibody is made against a synthetic peptide (trimethylated at Lys4) corresponding to amino acids 1-12 of human histone H3.
Modifications:
Methylation
Isotype:
IgG1κ
Species Reactivity:
Human
Vertebrates
Species Reactivity Note:
Human. The immunogen sequence is identical in a wide range of animal and plant species.
Application Notes:
Chromatin Immunoprecipitation:
Sonicated chromatin prepared from untreated or colcemid treated 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-trimethyl-Histone H3 (Lys4) antibody and the Magna ChIP G Kit (Cat. #17-611). Successful immunoprecipitation of trimethyl-Histone H3 (Lys4) associated DNA fragments was verified by qPCR using primers ampliflying a region of the human B-Globin promoter or using GAPDH promoter Control 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:
Representative lot data.
HeLa acid extract were resolved by electrophoresis, transferred to PVDF membrane and probed with Anti-trimethyl Histone H3 (Lys4), clone CMA304 at 1 μg/ml (lane 1) or 0.5 μg/ml (lane 2). Proteins were visualized using a goat anti-mouse secondary antibody conjugated to HRP (Cat. #AP124P) and a chemiluminescence detection system (Please see figures).
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Control:
Included negative control mouse IgG antibody and control primers specific for human GAPDH promoter.
Quality Assurance:
Chromatin Immunoprecipitation:
Sonicated chromatin prepared from colcemid-reated HeLa cells (1 X 106 cell equivalents pe |