Home
Back
Beer-Lambert Law:
| From: | To: |
The Beer-Lambert law relates the absorption of light to the properties of the material through which the light is traveling. It states that the absorbance of a solution is directly proportional to the concentration of the absorbing species and the path length of the light through the solution.
The calculator uses the Beer-Lambert law equation:
Where:
Explanation: The equation calculates protein concentration based on UV absorbance at 280 nm, which is primarily due to tryptophan and tyrosine residues in proteins.
Details: Accurate protein concentration measurement is essential for various biochemical applications including enzyme kinetics, protein purification, Western blotting, and protein-protein interaction studies.
Tips: Enter absorbance at 280 nm, extinction coefficient value, and path length. Ensure all values are positive numbers. The path length is typically 1.0 cm for standard cuvettes.
Q1: Why measure absorbance at 280 nm?
A: Proteins absorb light at 280 nm primarily due to their tryptophan and tyrosine content, making it a convenient method for protein quantification.
Q2: How do I determine the extinction coefficient?
A: The extinction coefficient can be calculated from the protein's amino acid sequence or measured experimentally using known protein concentrations.
Q3: What is a typical path length?
A: Standard cuvettes have a 1.0 cm path length, but microvolume spectrophotometers may use shorter path lengths.
Q4: Are there limitations to this method?
A: This method assumes the protein sample is pure and doesn't account for nucleic acid contamination, which also absorbs at 260 nm.
Q5: What if my sample is too concentrated?
A: For accurate measurements, absorbance values should be between 0.1 and 1.0. Dilute samples that exceed this range.