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Typically we determine the concentration of proteins using a 280 nm reading. However, it is reasonable to use 205 nm. I was curious about the effectiveness of this method.

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What evidence do you have for the reasonableness of reading at 205? –  MattDMo Mar 29 '13 at 22:13
    
@MattDMo: I guess it is because usually nucleic acids are measured at 260. However, I am not sure 205 would remove DNA background noise... These are very approximative quantifications anyways, and only give a general idea of protein concentration. –  nico Mar 30 '13 at 10:51
    
@MattDMo, I didn't want to lead the question but I was looking over this paper. There is a scientific basis for the quetion. onlinelibrary.wiley.com/doi/10.1002/pro.2253/abstract –  bobthejoe Apr 1 '13 at 23:59
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1 Answer 1

This may be what you are looking for. The conversion formula is:

$$ \text{Protein concentration (mg/ml)} = \frac{ (\text{Absorbance at 205 nm})} {31} $$

The first, Scopes, does indicate that nucleic acid contaminants will confounding 205 nm absorbance readings. It seems as though this spetrophotometic calculation is feasible for relatively pure protein species.

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DNA absorbance seems to be as high at 205 as at 260. faculty.evansville.edu/be6/b4454/DNAspectrum.gif?dur=2225 –  nico Mar 30 '13 at 10:58
    
You would appear to be correct. What is not shown is relative absorbance of protein and nucleic acid at 205 nm. NB: I have not followed up with the two references linked from that page I posted. –  leonardo Mar 30 '13 at 19:05
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I wanted to emphasize that A280nm is more sensitive, as tryptophan's absorbance is highest. So you can detect the smallest concentrations with A280nm. For proteins with little-to-no aromatics, or for proteins with atypical compositions (and unknown absorption coefficients), the absorbance per-unit-peptide bond is more general. (That's what A205nm is probing.) But as others have said, contaminating nucleic acids will also absorb at that wavelength. –  Ryan Apr 5 '13 at 4:24
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