Coomassie Blue protein assay - Interference from phenolic acids (Aug/16/2006 )
I've been asked to check for protein/amino acids in some aqueous samples. We use Coomassie blue to test protein/enzyme samples normally, but these samples also contain phenolics, such as Ferulic acid and vanillin. Does anyone know if these will interfere with the assay? If so, is there anything else you might suggest?
you can use bca or lowry.
keep in mind that none of these methods will detect single amino acids.
also, if you want to find out about interference, try it with the range of the contaminant that you expect to find.
An idea: you can check that If you have enough of your samples - percipitate, dialyse or ultrafiltrate proteins and use supernatant as the sample for protein assay.
PS. I think they may interfere, BCA may be better for this.
PS. I think they may interfere, BCA may be better for this.
I'm actually trying to find out if I have protein in my sample, because my PI told me to check (although I wasn't expecting any), but I'm most interested in the phenolics, so I'd want to keep that rather than the proteins, I'm not sure if that's possible with the above methods though.
I tried to check for interference with pure compounds, but we normally dissolve them up in 50:50 MeOH: H2O, which apparently has a similar reaction to 10 µg/ml BSA. They're not generally that soluble in H2O only, so it's a bit of a pain to check. I did run my samples anyway, just as a looksee and they seem to have less than 4µg/ml, so I think I'll have to check on the interference to be sure.
To try the other ones you guys suggested I'll need a little more info because I'm a chemist by training rather than a biochemist, so I'm not sure what you mean.
Thanks for your help
the methods mentioned in this thread all require you to sacrifice a (small, miniscule) portion of your sample. bca and lowry are destructive and coomassie will contaminate (and modify) the sample.
you could precipitate protein with acetone. if the phenolics are soluble in acetone then you may be able to purify them away from the proteins in this manner.