EZ-Magna ChIP™ A/G Chromatin Immunoprecipitation Kit

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 carried using the Magna ChIPTM A/G chromatin immunoprecipitation kit following the manufacturer's instructions (Merck Millipore, Watford, UK). In brief, ASM cells were grown in culture and treated as described above. Following stimulation the cells were formalin-fixed. The cells were sonicated to shear DNA (5 × 15 s at 40% amplitude) to ∼200–1000 bp in length. Brd4 and c-Myc were immunoprecipitated using 1 μg of antibody (Santa Cruz Biotechnology, Santa Cruz, CA). After reverse cross-linking, the DNA was purified and quantitative PCR carried out using SybrGreen (Qiagen) and the following primers: pCXCL8 forward, 5′-GGGCCATCAGTTGCAAATC-3′; pCXCL8 reverse, 5′-TCCTTCCGGTGGTTTCTTC-3′; pIL6 forward, 5′-CAAGCCTGGGATTATGAAGAAGG-3′; and pIL6 reverse, 5′-AGCACTGGCAGCACAAGGCAAAC-3′. The amount of DNA recovered by ChIP was normalized to an input control. IgG isotype antibodies (provided by Millipore) were used as a negative control for the ChIP assay.

Full paper   Login or join for free to view the full paper.

Reviews

Discussion

Papers

Check out relevant papers found by Labettor's AI that are relevant for performing ChIP Human - ASM using EZ-Magna ChIP™ A/G Chromatin Immunoprecipitation Kit from Merck Millipore.

Manufacturer protocol

Download the product protocol from Merck Millipore for EZ-Magna ChIP™ A/G Chromatin Immunoprecipitation Kit below.

Download manufacturer protocol

Videos

Check out videos that might be relevant for performing ChIP Human - ASM using EZ-Magna ChIP™ A/G Chromatin Immunoprecipitation Kit 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.