ChIPAb+Dimethyl-Histone H3 (Lys27);

Stock Code: 3586325
Manufacturer Part No: 17-10108

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Descriptions

Analysis Note

ControlIncludes negative control rabbit supernatant and primers specific for human β-globin promoter.

Application

Research Sub CategoryHistones

This ChIPAb+ Dimethyl-Histone H3 (Lys27) -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 either 2 µL of Negative Control Supernatant or 2 µL of Anti-dimethyl-Histone H3 (Lys27) and the Magna ChIP^® A Kit (Cat. # 17-610).
Successful immunoprecipitation of dimethyl-Histone H3 (Lys27) associated DNA fragments was verified by qPCR using ChIP Primers, human β-globin as a positive locus, and GAPDH promoter primers as a negative locus (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.
A 1:1000-1:5000 dilution of a previous lot detected dimethyl-Histone H3 in acid extracted proteins from HeLa cells, but did not detect unmethylated recombinant Histone H3 (Catalog # 14-494).
Recombinant Histone H3 (lane 1) and HeLa cell acid precipitate (lane 2) were resolved by electrophoresis, transferred to nitrocellulose and probed with anti-dimethyl-Histone H3 (Lys27) (1:1000 dilution).
Proteins were visualized using a goat anti-rabbit secondary antibody conjugated to HRP and a chemiluminescence detection system.
Arrow indicates dimethyl-histone H3 (~17 kDa) (Please see figures).

Peptide Inhibition Assay (PIA):
Representative lot data.
0.5-2 µM of histone H3 peptides containing dimethyl-Lys27 abolished detection of histone H3 by anti-dimethyl-Histone H3 (Lys27) (1:1000 dilution) in immunoblots of acid extracted proteins from HeLa cells.
Acid extracted proteins from HeLa cells were resolved by electrophoresis, transferred to nitrocellulose and probed with anti-dimethyl-Histone H3 (Lys27) (lane 1) or anti-dimethyl-Histone H3 (Lys27) preabsorbed with 0.5 mM of histone H3 peptides containing the following modifications:
Lane 2: dimethyl-lysine 23
Lane 3: dimethyl-lysine 27
Lane 4: dimethyl-lysine 9
A 1:1000 dilution of the primary antibody was used.
Proteins were visualized using a goat anti-rabbit secondary antibody conjugated to HRP and a chemiluminescence detection system.
Arrow indicates dimethyl-histone H3 (~17 kDa) (Please see figures).

Research CategoryEpigenetics & Nuclear Function

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