I have recently purified some IgG via affinity chromatography. As a result the elutions from my column were pretty dilute. I concentrated them by using PEG to pull out the excess water but I think the PEG leaked into my IgG prep. Now when I run a Bradford or BCA to assess total protein content I get very different results. I'm just not sure which assay to trust (if I can trust either of them at all!). Does anyone know how PEG affects the results of either of these two assays?
I never trust Bradford, I think it is more likely to interfere with Bradford's assay.
What do you use as a standard? is it the same for both assays?
reducing agents, metal chelators and some detergents are likely to interfere with Bradford, however I have no idea about PEG. BCA on the other hand is known to be more reliable and sensitive.
As a rule of thumb you tend to avoid any detergent when using Bradford, so I would distrust that result. PEG doesn't chelate copper nor is a reducing agent so your BCA results should be fine. I've heard that addition of cyclodextrins can alliviate the detergent problem in Bradford but these too can cause error, so I wouldn't bother and just use the BCA results. You could dilute your sample and run the BCA again, if it corresponds to your original result then it's unlikely PEG is causing issues.
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