Can ChIP be applied on tissue samples? - (Jul/13/2005 )
Hi, do you know wether the Chromatin Immunoprecipitation can be applied on the tissue samples? Theoretically, it seems applausible. Is there anybody with such experiences?
While I haven't tried it myself, I do know someone in another lab who tried it with frozen sections. It would seem as if there was success in amplifying the target sequence, but there was also additional trobleshooting that needed to be done before really trusting the result. I never did find out if the experiment truly worked or not.
There are alot of concerns in doing ChIP in tissue, however. The primary concern is how the tissue is obtained and what method was used to preserve it. Even if the tissue is fixed immediately, there's no guarantee the TF of interest is still bound to DNA. Over/under crosslinking becomes an issue as well. The large amount of structural material in tissue may interfere. Finally, the mechanical disruption of the tissue during grinding may impact your TF binding to DNA. It might be a tough experiment, but I do believe its been published before. I'll see if I can't dig up the reference somewhere.
Good luck and post back with progress
im also trying to do Chip on tumor tissues...following the protocol published in 2002 Methods journal. here is the link http://www.genomecenter.ucdavis.edu/farnha...ls/tissues.html
right now im standardizing the sonication step and i hope the experiment works.
what is the "preblocked Staph A cells"
They're using the whole cells instead of just proteinA/G.
Hi, all, I am so glad to hear you all use tissues to do ChIP. I am working on mouse tissues for a few months till now. I am not feel lonely now. I stored tissues into -80. It seems work well. I still don't know how long it can be stored in -80 and not effect ChIP.
I use antibody from Upstate, one of them worked well. Another one not. I wrote to Upstate, then found that I made a mistake. The other antibody is not recommend for ChIP. I am looking for ChIP grade antobdy now.
To me, when I use the same sonication condition. The tissue stored in -80 is more easy to be sonicated than fresh tissue, but the fresh tissue is easy to do ChIP.
I am still testing the results. There is a long way .......