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Calculations for Protein A280 Measurements
The Protein A280 application uses the
Beer-Lambert equation
absorbance with concentration. Solving
Beer's law for concentration yields the
equation at the right.
The extinction coefficient of a peptide or
protein is related to its tryptophan (W),
tyrosin (Y) and cysteine (C) amino acid
composition.
Tip: The extinction coefficient is
wavelength specific for each protein and
can be affected by buffer type, ionic
strength and pH.
Thermo Scientific
to correlate
Beer-Lambert Equation (solved for concentration)
c = A / (
* b)
where:
A = UV absorbance in absorbance units (AU)
= wavelength-dependent molar absorptivity coefficient (or
extinction coefficient) in liter/mol-cm
b = pathlength in cm
c = analyte concentration in moles/liter or molarity (M)
Note: Dividing the measured absorbance of a sample
solution by its molar extinction coefficient yields the molar
concentration of the sample. See
Coefficients
for more information regarding molar vs. mass
concentration values.
Extinction Coefficients for Proteins
At 280 nm, the extinction coefficient is approximated by the
weighted sum of the 280 nm molar extinction coefficients of
the three constituent amino acids, as described in this
equation:
= (nW * 5500) + (nY * 1490) + (nC * 125)
where:
= molar extinction coefficient
n
= number of each amino acid residue
5500, 1490 and 125
= amino acid molar absorptivities at
280 nm
5
Protein Applications
Measure Protein A280
Published Extinction
NanoDrop One User Guide
73
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