1a f, Mm a – Eppendorf BioSpectrometer basic User Manual
Page 95
95
Evaluation procedure
Eppendorf BioSpectrometer
®
basic
English (EN)
12.5.4
Calculating the factor for protein in "General Method Parameter"
This section only covers the calculation of the protein components in the
Dye labels and Proteins direct
UV method groups. For these method groups, the protein component is selected in the parameters (see
Method parameters on p. 35). The protein component is assigned a factor that will be entered in the
General Method Parameter/Proteins function for each protein. Alternatively, A
0.1%
or the absorbance
coefficient plus the molar mass of the protein can be entered instead of the factor. In this case, the factor is
calculated as follows:
F = factor for the protein; unit: g/L.
A
0.1%
= absorbance of the protein at a concentration of 0.1
% (1 g/L).
When entering the molar absorbance coefficient and the relative molar mass of the protein
A
0.1%
can be
calculated on this basis:
ε
P
= molar absorbance coefficient of the protein; unit: cm
-1
M
-1
.
MM
P
= relative molar mass of the protein; unit: Da (entry in
General Method Parameter in kDa).
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. 91). 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
• For
dsDNA the calculation of the molar concentration is based on the assumption of a
double-stranded nucleic acid. For the
ssDNA, RNA and Oligo methods, a single-stranded
nucleic acid is assumed.
• For methods which have been reprogrammed via
group,
Nucleic acids method group, always double-stranded nucleic acids are assumed
for calculating the molar concentration.
%
1
.
0
1
A
F
P
P
P
MM
A
H
%
1
.
0
Dye
Dye
F
H
6
10