41Can UV absorbance distinguish dsDNA, ssDNA and RNA?42Why does RNA contamination lead to an overestimate of DNA concentration?43How do absorbance and fluorescence methods differ?44Can purity ratios indicate nucleic acid integrity?45Why can proteins be quantified directly using A280?46Can all proteins use the same A280 conversion factor?47What are the limitations of direct A280 quantification?48What is the basic principle of the BCA assay?49What commonly interferes with the BCA assay?50What is the basic principle of the Bradford assay?

What are the limitations of direct A280 quantification?

Direct A280 quantification is useful when the extinction coefficient is known and background interference is controlled. Different aromatic-residue and disulfide contents mean equal mass concentrations need not produce equal absorbance.

Confirm protein identity and coefficient units, then check the buffer and other components for UV absorption. A generic coefficient provides only an approximation for unknown mixtures or crude extracts.

If using a colorimetric assay, check reagent compatibility and differences between standard and sample protein responses, and calculate within the valid calibration range. Changing methods does not automatically eliminate every bias.