Eppendorf BioSpectrometer kinetic - Manual (Page 96)

Evaluation procedure
Eppendorf BioSpectrometer
® kinetic
English (EN)
96
12.6 Special evaluation procedures for the dye methods
12.6.1 Calculating the factor for the dye from the absorbance coefficient
For the dye methods the concentration of the dye is calculated using a factor from the measured
absorbance (see Evaluation with factor or standard on p. 90). The factor is entered for each dye in the
General Method Parameter/Dyes function. Alternatively, you can enter the absorbance coefficient. In this
case, the factor is calculated as follows:
F = factor for the dye; unit: pmol/μL.
ε = absorbance coefficient for the dye, unit: cm -1 M -1
12.6.2 Calculation of the FOI
As a value for the ratio of dye molecules to the number of nucleotides in the nucleic acid the frequency of
incorporation (FOI) is calculated and displayed for the dye methods. The calculation can be selected for two
different result units:
MOLECULE dye/kb unit
pmole/μg DNA (or RNA) unit
A
YYY
= absorbance of the dye.
A
XXX
= absorbance of the nucleic acid.
MM
nt
= average molar mass of the nucleotides: 330 g/mol.
F
NA
= factor for calculating the nucleic acid
ε
dye
= absorbance coefficient for the dye, unit: cm -1 M -1
Dye
Dye
F
H
6
10
NA XXX
nt
Dye
YYY
F A
MM A
FOI
u
u
u
6 10
H
NA XXX Dye
YYY
F A
A
FOI
u
u
9 10
H