Hello
I would like to do Co-Immunoprecipitation of two proteins which I would like to detect by Western Blot. I read that the technique is not quite easy. The lysis buffer (especially the detergents) is a critical parameter. Actually I am not sure if my proteins have an interaction. So which control (two proteins which interact definitely) could I use to establish the Co-Ip protocol to exclude that the procedure is responsible for the failure. Has anyone a idea?
It would be very helpful for me.
Thanks
Tiffy

You could use any known multimeric protein complex for which there are commercially available antibodies that you can afford.

e.g. you could Co-ip a potassium channel alpha subunit with its beta subunit antibody
You could Co-IP an NMDA subunit with PSD 95

Use pubmed to look up two proteins known to interact and which express in the cell type you are using for experiments, and for which there are commercially available antibodies. Alternatively look up in Pubmed and pair of proteins known to interact, preferably the same size and with similar molecular structure to your proteins of interest and express them transiently. There are plenty of epitope tagged proteins tat you can request from other labs or that you can buy the plasmids for which you can express and use to pull down a binding partner.

This kit even comes with a positive control lysate

http://www.piercenet.com/Products/Browse.cfm?fldID=9C471132-0F72-4F39-8DF0-455FB515718F

Dear Tiffy.
The question that you asked is one that many of us whom are doing CO-IP are strageling each time we are starting doing a new COIP experiment.
If you ask me there isnt any two candidates that you should use in order to calibrate your system .
Choosing the correct lysis buffer is very esential step and depend upon the proteins and their compexes. The protocol is easy to do but needs to calibrate with the proteins that you are interesed in. Dont try with ones that are not in your research.
First try to do a regular IP with the protein of your intrest with several different mild lysis buffers. Usualy I use 20mMTris and 150mM NaCl pH 7.4 with 0.1%NP40. you can play with the NP40 % 0.05 up to 0.3. Find out which Ab to pull down your protein depending upon the othe protein you want to clook at the co-ip step. mono for IP and poly for WB or vise veres.
So that is the first step. After you find out which one is the best go to the secound step COIP.
Guy