NanoPhotometer
®
NP80/N60/N50/C40
User Guide Version 3.1
48
initiate the measurement.
C
ALCULATIONS
Nucleic Acid Concentration
To determine the concentration of nucleic acids in solution, the absorbance as measured at the
wavelength of 260 nm is used. The function describing the relationship between concentration
and absorbance is a modification of the Beer-Lambert law equation. The concentration of the
nucleic acid samples can be calculated with or without background correction depending on
enabled/disabled background correction option.
Without background correction:
C = A
260
*
ε
nuc
* Đ
With background correction:
C =
(
A
260
- A
BKG
)
*
ε
nuc
*
Đ
C
Concentration in ng/µl
A
260
Absorbance at 260 nm (10 mm path)
A
BKG
Absorbance at selected background wavelength (10 mm path)
Đ
Manual dilution factor
ε
nuc
Extinction coefficient/nucleic acid factor in ng*cm/µl
Table 1. Nucleic acids extinction coefficients (
ε
nuc
)
Type
ε
nuc
dsDNA
50 ng*cm/µl
ssDNA
37 ng*cm/µl
RNA
40 ng*cm/µl
miRNA
33 ng*cm/µl
Oligo
33 ng*cm/µl
miRNA Seq. calculated via extinction coefficient of constituent nucleotides entered
Oligo Seq.
calculated via extinction coefficient of constituent nucleotides entered
Custom
Option to enter any factor between 15 and 150 ng*cm/µl
Dye-labeled Nucleic Acid Concentration
For dye-labeled nucleic acids, the concentration of the nucleic acid is calculated using a
modified form of the Beer-Lambert equation. For these calculations, the instrument considers
the absorption maximum of the dye and a certain dye-specific correction factor at 260 nm (see
Table 2). The concentration of a dye-labeled nucleic acid is calculated with or without
background/dye correction as follows:
