Assessment of Nucleic Acid Yield/Quality
Molecular Diagnostics
(MDx) Review 550
METHOD EXPLANATION ASSESSMENT
Yield
Purity
DNA μg/mL: A260 × 50
RNA μg/mL: A260 × 40
Brighter bands = higher yield
DNA A260/A280 ratio:
1.6–2.0 = good quality
<1.6 = protein contamination, speci-
men must be reprocessed
>2 = possible contamination with RNA
RNA A260/A280 ratio:
≥2 = good quality
DNA:
high MW fragments = good quality
RNA 28S/18S ratio:
2 = good quality
<2 = smaller fragments due to RNase
degradation
Spectrophotometry: Absorbance
@ 260 nm
Gel electrophoresis/densitometry
Spectrophotometry: Absorbance
@ 260 nm/absorbance
@ 280 nm
Gel electrophoresis/densitometry
Nucleic acids absorb light at
260 nm.
Intensity of bands is proportional
to nucleic acid concentration.
Compare to standard.
Nucleic acids absorb light at
260 nm. Absorption peak for
protein is 280 nm.
Position of bands is related to
MW. Lower MW fragments
migrate furthest from point
of application.
