mysterious thing! - (Nov/30/2008 )

Hi,
I am facing with the mysterious thing. When the OD that I get with Brad ford assay is less than one for the protein lysate, my western blot dies not work, while in the same membrane more concentrated peorein attaed and I get band for WB. Although for both samples I added the same amounth of protein!
Has any on can explain that and give me some sugestion. What should I do with this matter? Even for the actin I didnt get any band.

thanks,
Sara

-saraarasus-

Degraded sample? I think Bradford works by binding to specific combinations of residues, if your sample is degraded there may be less binding capacity and thus an anomalous result.

-bob1-

Hi, Thank for your reply. But this problem has happened before too. When the protein concentration is less than certain amount WB doesnt work although I try to load enough protein.
I am not sure maybe they pass through !!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!

QUOTE (bob1 @ Nov 30 2008, 04:01 PM)

Degraded sample? I think Bradford works by binding to specific combinations of residues, if your sample is degraded there may be less binding capacity and thus an anomalous result.

-saraarasus-