ChIP assay - (Jun/04/2004 )
has anyone here performed a ChIp assay on myocytes or heart tissues?
I am using the upstate protocol in these systems but it is not working. does anyone had success in these system? if yes can u share the protocol and relevant advice.
I have done some ChIPs with cultured cells using Upstate ChIP kit and anti-acetylated histone antibodies. It gave me satisfactory results at first try. I have studied a number of loci with primers spanning about 200 bp. Some loci may not easy to amplify after ChIP. I think the critical step is DNA sonication which should break your DNA to 200-1000 bp fragments, not less and not more. You have to do a preliminary experiment to optimize the sonicating settings. Apart from that, the rest is easy to follow.
Hope it helps.
Farnham PJ has published a number of protocols in his papers and they may be of some use. We are currently trying his protocol (just started)
Hope it works