Description
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 H4 set includes the Histone H4 antibody, a negative control antibody (normal rabbit IgG), and qPCR primers which amplify a 110 bp region of human β-Globin. The Histone H4 and negative control antibodies are supplied in a scalable "per ChIP" reaction size and can be used to functionally validate the precipitation of Histone H4-associated chromatin.
~11 kDa
Immunogen
Epitope: a.a 25-28
KLH-conjugated synthetic peptide corres-ponding to amino acids 17-28 of Histone H4.
Application
Chromatin Immunoprecipitation:
Sonicated chromatin prepared from HeLa cells (1 X 106 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 2 µL of either normal rabbit IgG or 2 µL of Anti-Histone H4 antibody and the Magna ChIP A Kit (Cat. # 17-610).
Successful immunoprecipitation of total Histone H4 associated DNA fragments was verified by qPCR using ChIP Primers, β-Globin, as well as 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 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 3), HeLa Acid Extract from sodium butyrate treated cells (Lane 2), HeLa Acid Extract from colcemid treated cells (Lane 4) and recombinant histone H4 (Lane 5) were resolved by electrophoresis, transferred to PVDF membranes and probed with Anti-Histone H4 (1:1,000 dilution).
Proteins were visualized using a Donkey anti-rabbit IgG conjugated to HRP and visualized using a chemiluminescence detection system. (Please see figures).
This ChIPAb+ Histone H4 -ChIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.
Biochem/physiol Actions
Wide range of cross-reactivity expected based on sequence homology.
Physical form
Anti-Histone H4 (rabbit monoclonal IgG). One vial containing 50 µL of protein A purified rabbit monoclonal IgG supernatant in buffer containing 0.1 M Tris-Glycine (pH 7.4), 150 mM NaCl with 0.05% sodium azide before addition of glycerol to 40%. Store at -20°C.
Normal Rabbit IgG. One vial containing 75 µg of normal rabbit IgG in 75 µL storage buffer. Store at -20°C.
ChIP Primers, β-Globin. One vial containing 75 μL of 5 μM of each primer specific for human β-Globin. Store at -20°C.
FOR: AGG ACA GGT ACG GCT GTC ATC
REV: TTT ATG CCC AGC CCT GGC TC
Format: Purified
Analysis Note
Chromatin Immunoprecipitation:
Sonicated chromatin prepared from HeLa cells (1 X 106 cell equivalents per IP) were subjected to chromatin immunoprecipitation using 2 µL of either normal rabbit IgG or 2 µL of Anti-Histone H4 antibody and the Magna ChIP® A Kit (Cat. #17-610). Successful immunoprecipitation of total Histone H4 associated DNA fragments was verified by qPCR using ChIP Primers, β-Globin (Please see figures).
Please refer to the EZ-Magna ChIP A (Cat. # 17-408) or EZ-ChIP (Cat. # 17-371) protocol for experimental details.
Control
Includes negative control antibody (normal rabbit IgG) and primers specific for human β-Globin.
Legal Information
MAGNA CHIP is a registered trademark of Merck KGaA, Darmstadt, Germany
UPSTATE is a registered trademark of Merck KGaA, Darmstadt, Germany
