Hi there,

This is not necessary because the time required for protein determination is short and also samples are denatured during the process so possible protease action will be negligeable.

Dear @Botho

i agree with Dominique that the inibithor is not certanilly requested for sample stability during the BCA asay.

However, I think that to check if this compunds may interfeere with the BCA assay you can compare the absorbance of a blank contaning the inihibotrs and with a blank with out the inibithorin the BCA assay. In this way you will be sure that the inibhitor is not able to reduce the cu(II) to Cu(I) and not produce a background signal in the BCA assay.

ciao

Manuele

Hi

On the other hand, you should check if you have EDTA among the inhibitors you use because it negatively interferes with the test.

First, your proteinase inhibitor is a protein (correct?) so it will have an effect on protein concentration. Your standards are BSA dilutions, which are also proteins, so you don't need the proteinase inhibitor here; proteins are proteins. But you do need a blank with lysis buffer plus the same amount of proteinase inhibitor as you have in the testing samples to run in your BCA assay; the concentration of this lysis buffer/proteinase inhibitor mix will be subtracted from the concentration of your samples/lysis buffer/proteinase inhbitors mix when you get the BCA results.

(your protein/lysis buffer/proteinase inhibitor) - (lysis buffer/proteinase inhibitor) = [your protein concentration]

If you are making a dilution curve of your protein/lysis buffer/proteinase inhibitor, make dilutions that include different concentrations of your sample and the blank described above (lysis buffer/proteinase inhibitor). A 1:1 dilution would give you a 50% concentration of your protein, etc. and the proteinase inhibitor will be constant in all the tubes.