While I was conducting the Bradford Assay experiment to determine the concentration of my unknowns, I ended up by having an A595 for my unknown samples that is higher than the A595 of the standards. How can I resolve this issue? wishes.. Micheal.

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    $\begingroup$ Just find out slop of your standard curve and then calculate unknown concentration from that. this might be helpful to you. $\endgroup$
    – Dexter
    Commented Oct 13, 2015 at 3:41
  • $\begingroup$ When you run a bradford assay the bound form of the dye shifts the absorption max to 595nm. This is what your spectrometer should be set to when you're running the assay. You make a standard graph because you can calculate an extinction coefficient ε from the slope of the graph. Beer's law says that A = εcl where your path length is determined by the cuvette you used, and c is your concentration. So you take your absorption that you obtained and solve for c! Canadianer makes an excellent point below, however, so make sure your methods make sense! $\endgroup$
    – CKM
    Commented Oct 13, 2015 at 19:31

1 Answer 1


You could, depending on what you are doing, extrapolate your standard curve to include the absorbance of your samples. This, however, is not recommended since absorbance is linear only within a certain range. It's best to dilute your samples so that they fall within the range of the standard curve while also ensuring that your standard curve is, in fact, linear.


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