Measure Protein A280
Thermo Scientific NanoDrop One User Guide 65
Calculations for Protein A280 Measurements
The Protein A280 application uses the Beer-Lambert
equation to correlate absorbance with concentration.
Solving Beer’s law for concentration yields the equation
at the right.
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
Published Extinction Coefficients for more information regarding molar
vs. mass concentration values.
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.
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
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