Bio-rad and ELISA - (Jun/22/2014 )

I'm just preparing for the exam and i found a question that i'm not sure about the answer

The concentration of same protein is determined by both Bio-rad and ELISA (which uses specific antibody against our protein of interest).

It's measured at 405nm (Bio-rad) and at 595nm (ELISA)

the concentrations obtained from Bio-rad are higher than those from ELISA method. And what could be the answer for such a difference?

Is it only that the choice of wavelegth is different, so the sample at 405nm is absorbing and therfore transmitting more light, which results in higher absorbance?

Or am i missing something?

BioRad is a company - which of their tests/equipment was used?

ah that's how our protocol was made, so that's how we call it between each other, sorry :)

Bio-rad protein assay

a couple of things:

the bio-rad protein assay is read at 595, that means that the elisa is probably the one read at 405 (or you read them wrong and that explains the variability).

the elisa is measuring a specific protein (and is probably not measuring 100%, but that's another issue).

the bio-rad measures all proteins (and, maybe, some buffer components) present.

also, accuracy of the bio-rad depends on the proteins present in solution and the protein used to generate the standard curve (there's protein to protein variability in dye-binding protein assays).

great that's what i thought as well, thanks :)