Hi,
I am doing WB with heart tissues (Rat) these days and having a problem of high background. We have no issue of getting high background for WB done with cells or other tissues, the protocol is well established.
Any suggestions?
Thank you
What is your method of detection? In my experience heart tissue may cause background issues when using fluorescent secondary antibodies. Never had any problems using chemiluminescent based detection.
Hi Yvette,
:) thanks for the info.. Yes we did longer blocking.. but now the target band masks. We tried to increase the protein loaded/well. It gives a huge nonspecific band with a very faint target band.
Cheers,
Susara
Blocking with BSA always gives sharp bands compared to skimmed milk. You can try blocking with 5-10% BSA in TBST for 1-2 hours.
Remember to filter the BSA after preparation.
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