Posted 30 July 2011 - 02:12 PM
I am stuck in optimizing an antibody which gives a LOT OF background to get the endogenous protein. it is supernatant so I cannot dilute so much. Do you have any idea how to remove all those non specific bindings?
So far, I have tried the following and none of them worked properly:
1-different blocking agents: 5% milk, 3% and 5% BSA, mix of BSA and Milk - the best one was 5% BSA overnight
2- small dilutions with 10% BSA - no work
3- I stripped the blot once, it didnot help
It drives me crazy. please help
Posted 30 July 2011 - 02:57 PM
Probably you're used o much antibodies in your solutions.
Posted 12 August 2011 - 03:05 AM