EZ-ChIP™

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	-Just because an antibody works well in a Western blot does not always indicate it will perform well in chromatin immunoprecipitation. Unlike a Western blot that detects proteins that have been denatured, a ChIP antibody must recognize the target protein in its native state. Use ChIP-validated antibody. -Use 2-10 μg of your ChIP antibody depending on the abundance of your protein target.

Protocol tips
-Just because an antibody works well in a Western blot does not always indicate it will perform well in chromatin immunoprecipitation. Unlike a Western blot that detects proteins that have been denatured, a ChIP antibody must recognize the target protein in its native state. Use ChIP-validated antibody. -Use 2-10 μg of your ChIP antibody depending on the abundance of your protein target.

Publication protocol

ChIP analysis was performed using EZ ChIP (Upstate Biotechnology, Lake Placid, NY, USA) according to the manufacturer’s protocols with certain modifications. Anti-polymerase II (Pol II), anti-H3Ac and anti-H3K9me2 antibodies were obtained from Upstate Biotechnology. Anti-H3K4me3, anti-H3K27me3 and anti-KDM6b were obtained from Abcam (Cambridge, UK). Anti-BMI-1 was obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). DNA fragments were amplified using PCR. The quantitative RT-PCR values were normalized by that of the corresponding input. The primer set that was used in the PCR is as follows: Nkcc1 (AF_051561) Forward 5′-AGGAGTCCACAGGGTCCGTGC-3′, Reverse 5′-CCCACTAAGGAGCCGGGGGAG-3′.

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Reviews

Discussion

Papers

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Manufacturer protocol

Download the product protocol from Merck Millipore for EZ-ChIP™ below.

Download manufacturer protocol

Videos

Check out videos that might be relevant for performing ChIP Rat - Mesenteric arteries using EZ-ChIP™ from Merck Millipore. Please note that these videos are representative and steps or experiment specific processes must be kept in mind to expect desired results.