Articles
DNA Quantification (Absorbance Mode)
August 30, 2006
•Application Note: DNA Quantification (Absorbance Mode)
The most common method for quantifying DNA samples is by conventional absorbance measurements; nucleic acids have an absorption maximum at 260nm. Most samples contain contaminates such as proteins and single stranded DNA/RNA that absorb maximally at 280nm. The equation for calculating DNA in the presence of contaminates is:
The higher the ratio, the more pure the DNA sample. It is acceptable to have a ratio between 1.8 and 2.0 for a cuvette spectrophotometer.
Comparing Results of a Spectrophometer and a Microplate Reader Absorbance is defined by Beer-Lambert equation
where e= molar coefficient, b= pathlength and c=concentration. When the molar coefficient and pathlength are constant, absorbance is proportional to the concentration.
For a standard cuvette reader, the pathlength is usually defined as 1 centimeter. Therefore, with a conventional absorbance reading an A260 of 0.1 O.D. corresponds to 5µg/ml dsDNA solution. Because of the shorter pathlength in a microplate reader, this value is somewhat smaller (0.07 O.D. corresponds to 5µg/ml dsDNA solution).
Click Here To Download:
•Application Note: DNA Quantification (Absorbance Mode)
SOURCE: BMG LABTECH, GmbH
