The elimination of ribonucleic acid interference in the spectrophotometric determination of protein concentration

Abstract

A simple, sensitive and rapid spectrophotometric method for the determination of protein concentration in the presence of RNA contaminants is described. Only small spectral differences exist between RNAs from different sources and to eliminate their interference, two wavelengths must be selected where absorbance due to RNA is identical. The isoabsorbance wavelengths must lie between 235 nm and 226 nm, the region where with decrease in wavelength the increase in absorbance is linear for most proteins. From data presented wavelengths of 233,0 nm and 226,0 nm were selected as the most suitable. A linear relationship between difference in absorbance (226, 0 nm-233, 0 nm) and protein concentration was observed for pure protein preparations up to 300 µg/ml and for protein: RNA mixtures. Protein concentrations determined by this method were comparable with those determined by the colorimetric method of Lowry. Differences in isoabsorbance wavelengths selected in this study and those reported by other workers are discussed.