Could I denature the proteins that were transferred on a membrane?
Posted 07 October 2009 - 01:13 AM
Posted 07 October 2009 - 05:08 AM
Posted 08 October 2009 - 02:33 AM
Thanks for your suggestion. But I think if the internal control is carried out separately, it would not be a loading control for my target protein. They should be performed on the same lane.
Interesting problem... Assuming there's not a better antibody for beta-actin, could you load two samples before running and transferring the gel -- one that is boiled and reduced, and another that is not?
Today I tried a high stringent condion. Wasing the membrane with 8 M urea-PBST plus 10% beta-ME for 1 hr, the result is no signal detected. It seems that all the proteins shed away.
Maybe it is impossible to do that. To denature proteins may also disrupt the binding between proteins and a membrane.
I will look for another antibody working well on native samples.
Posted 08 October 2009 - 06:52 AM