Analysis Note

ControlIncludes normal rabbit serum and primers specific for human β-globin.

Application

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

Chromatin Immunoprecipitation:
Representative lot data.
Sonicated chromatin prepared from HeLa cells (1 X 10E6 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 1 µL of either Normal Rabbit Serum,or 1 µL of Anti-Histone H3 (C-Term) and the Magna ChIP™ A Kit (Cat. # 17-610). Successful immunoprecipitation of histone H3 associated DNA fragments was verified by qPCR using ChIP Primers, human β-globin as a positive locus, and GAPDH promoter primers (22-004) as a negative locus. (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.
Acid extracted proteins from HeLa cells untreated (lane 1) or treated with sodium butyrate (lane 2) or colcemid (lane 3) and recombinant Histone H3 (lane 4) were resolved by electrophoresis, transferred to nitrocellulose and probed with anti-Histone H3, CT, pan (1:50,000 dilution). Proteins were visualized using a goat anti-rabbit secondary antibody conjugated to HRP and a chemiluminescence detection system.
Arrow indicates Histone H3 (~17 kDa). (Figure 3).

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.
The ChIPAb+ Histone H3 (C-Term) set includes the Histone H3 (C-Term) antibody, a Normal Rabbit Serum, and control primers which amplify a 110 bp region of ChIP Primers, human β-globin. The Histone H3 (C-Term) and negative controls are supplied in a scalable "per ChIP" reaction size and can be used to functionally validate the precipitation of Histone H3 (C-Term)-associated chromatin.

Immunogen

KLH-conjugated, synthetic peptide corresponding to the C-terminus of human Histone H3.

Legal Information

MAGNA CHIP is a registered trademark of Merck KGaA, Darmstadt, Germany

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

Packaging

25 assays per set. Recommended use: 1 µL of antibody per chromatin immunoprecipitation (dependent upon biological context).

Physical form

Anti-Histone H3 (C-Term) (rabbit polyclonal). One vial containing 25 µL of rabbit antiserum diluted 1:2 in storage buffer (0.02 M Phosphate, 0.25 M NaCl, 0.1% sodium azide) before the addition of glycerol to 30%. Store at -20°C.
Normal Rabbit Serum. One vial containing 25 µL of antiserum containing 0.05% sodium azide. Store at -20°C.
ChIP Primers, human β-globin. One vial containing 75 µL of 5 µM of each primer specific for the human β-globin promoter. Store at -20°C.
FOR: AGG ACA GGT ACG GCT GTC ATC
REV: TTT ATG CCC AGC CCT GGC TC

Quality

Chromatin Immunoprecipitation:
Representative lot data.
Sonicated chromatin prepared from HeLa cells (1 X 10E6 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 1 µL of either Normal Rabbit Serum, or 1 µL of Anti-Histone H3 (C-Term) and the Magna ChIP^® A Kit (Cat. # 17-610). Successful immunoprecipitation of histone H3 associated DNA fragments was verified by qPCR using ChIP Primers, human β-globin (Figure 1).
Please refer to the EZ-Magna ChIP™ A (Cat. # 17-408) or EZ-ChIP™ (Cat. # 17-371) protocol for experimental details.

Specificity

Recognizes C-Terminal region of Histone H3, Mr 17 kDa

Broad species cross-reactivity expected due to sequence homology.

Target description

Approx. 17 kDa