1. What is the Beer-Lambert Law?

The Beer-Lambert law describes the relationship between the absorption of light and the properties of the material through which the light is traveling. It is fundamental for determining protein concentrations in biochemical assays.

2. How Does the Calculator Work?

The calculator uses the Beer-Lambert law:

Where:

• \( c \) — Protein concentration (mg/mL)
• \( A \) — Absorbance (AU)
• \( \varepsilon \) — Extinction coefficient (M⁻¹ cm⁻¹)
• \( l \) — Path length (cm)

Explanation: The equation calculates protein concentration based on light absorption properties, where absorbance is directly proportional to concentration and path length.

3. Importance of Protein Concentration Measurement

Details: Accurate protein concentration determination is essential for biochemical experiments, protein purification, enzyme kinetics studies, and ensuring proper protein loading in various assays.

4. Using the Calculator

Tips: Enter absorbance value, extinction coefficient, and path length. Standard cuvettes typically have 1 cm path length. Ensure all values are positive and extinction coefficient is appropriate for your protein.

5. Frequently Asked Questions (FAQ)

Q1: What is a typical extinction coefficient for proteins?
A: Extinction coefficients vary by protein. Common values range from 10,000 to 50,000 M⁻¹ cm⁻¹. BSA has ε = 43,824 M⁻¹ cm⁻¹ at 280 nm.

Q2: Why measure at 280 nm?
A: Proteins absorb at 280 nm due to tryptophan and tyrosine residues. This wavelength is commonly used for protein concentration determination.

Q3: What affects absorbance accuracy?
A: Sample turbidity, buffer composition, protein aggregation, and proper blank correction can all affect absorbance measurements.

Q4: Can I use this for any protein?
A: This method works best for proteins with known extinction coefficients. For unknown proteins, other methods like Bradford or BCA assay may be preferred.

Q5: What is the linear range for absorbance measurements?
A: Typically 0.1 to 1.0 AU for most spectrophotometers. Values outside this range may not follow the Beer-Lambert law accurately.