NanoPhotometer
®
NP80/N60/N50/C40
User Guide Version 3.1
59
maximum of the dye, and a dye-specific extinction coefficient (see Table 4). The dye
concentration is calculated with or without background correction for are as follows:
With background correction:
C =
�
(A
max,dye
−
A
BKG
) * Ð
�
ε
dye
* 10
6
Without background correction:
C =
�
A
max,dye
* Ð
�
ε
dye
* 10
6
C
Concentration
A
max, dye
Absorbance at the max dye absorbance value (10 mm path)
A
BKG
Absorbance at selected background wavelength (10 mm path)
ε
dye
Extinction coefficient of dye in M
-1
*cm
-1
Ð
Dilution factor
Degree of Labeling (DOL)
DOL is the degree of labeling based on the average number of dye molecules coupled to a
protein molecule. The degree of labeling can be determined from the absorption spectrum of the
labeled antibody with or without background/dye correction as follows:
With background and with dye correction:
DOL =
(A
max, dye
−
A
BKG
) *
ε
prot
�
A
280
−
A
BKG
−
cf
dye
* (A
max, dye
−
A
BKG
)
� ∗ ε
dye
With background and w/o dye correction:
DOL =
(A
max, dye
−
A
BKG
*
ε
prot
(A
280
−
A
BKG
) *
ε
dye
W/o background and with dye correction:
DOL =
A
max, dye
*
ε
prot
�
A
280
−
cf
dye
* A
max, dye
�
*
ε
dye
W/o background and w/o dye correction:
DOL =
A
max, dye
*
ε
prot
A
280
*
ε
dye
