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Position: Discussion forums > Bioscience biotechniques discussion forums > Proteomics and protein biochemistry forum
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Problem about the minimum amount of protein to do immunoprecipitation
Posted by: formore (IP Hidden, New member, 1)
Date: January 20, 2005 09:17AM
Hi, I am a new one to do co-immunoprecipitation. I want to use it to do Kinase Activity Assay.
That is to say, I want to coprecipitate a kind of protein kinase from mouse embryo tissue extract. After the immunoprecipitation, the activity of the kinase would be detected by radioactive assay. The problem is, when I got several protocols of such immunoprecipitation, I found that most of them require the minimun amount of protein for the immunoprecipitation is 200 ug (protocol for mouse skeletal muscle) or 0.5-1mg (protocol for whole-cell extract on some book). Is that true? My embryo sample may only have the protein concentation of 1-2 ug/ul and 20-30 ug altogether. Since the latter radioactive assay should be very sensitive, I just wonder how could I handle this small protein sample for immunoprecipitation? Have you any idea about that? Would you please give me some suggestions? Thank you very much. yichaowu@hotmail.com
minimum protein quantity
Posted by: huangz123 (IP Hidden, Junior member, 19)
Date: January 22, 2005 03:32PM
It depends on what kinase assay you are going to perform. Normally it requres at least 200 ug protein in most IP protocol. However if your protein to be pulled down is abundant it may require less quantity. For example, 50ug of protein would be sufficient to do a cdk2 kinase assay from highly proliferative cells. Hope this helps.
Zhong
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