Beer-Lambert Protein Concentration Calculator
Determine protein concentration in solution using UV absorbance measurements, extinction coefficients, and optical path length.
Calculation Parameters
Specify your stoichiometry inputs.
Calculated Result
Expert Tip
The standard optical path length for cuvettes in spectrophotometers is exactly 1.0 cm.
How this Calculator Works
This calculator computes protein concentrations using the Beer-Lambert law. Select a preset or adjust parameters using the sliders:
- Absorbance (A): The measured UV intensity reduction at 280 nm (Default: 0.5)
- Extinction Coefficient (ε): Molar absorptivity of the protein in M⁻¹cm⁻¹ (Default: 43,824 for BSA)
- Path Length (b): Cuvette light path width in cm (Default: 1.0 cm)
- Molecular Weight (MW): Grams per mole of protein to calculate mass concentration (Default: 66,463 g/mol)
The results are computed instantly and updated in the results panel on the right.
Formula & Methodology
Where:
- c: Molar concentration of protein (mol/L or M).
- A: UV optical absorbance.
- ε: Extinction coefficient (M⁻¹cm⁻¹).
- b: Light path length (cm).
- MW: Protein molecular weight (g/mol).
Step-by-Step Calculation Example
To calculate concentration manually using BSA defaults, follow these steps:
- Identify the input parameters. For example:
- Absorbance (A) = 0.500
- Extinction (ε) = 43,824 M⁻¹cm⁻¹
- Path Length (b) = 1.0 cm
- MW = 66,463 g/mol
- Apply the formula:
1. Molar Concentration c = 0.500 / (43,824 * 1.0) = 1.141e-5 M 2. Micromolar c = 1.141e-5 * 1,000,000 = 11.41 µM 3. Mass Concentration = 1.141e-5 * 66,463 = 0.758 mg/mL
- Verify the calculated value which is updated instantly in the results panel on the right.
Frequently Asked Questions
Why is protein absorbance measured at 280 nm (A280)? ▼
A280 measures UV absorption by aromatic amino acid side chains, primarily Tryptophan (Trp) and Tyrosine (Tyr).
What is the formula for protein concentration using A280? ▼
Concentration (M) = A280 / (ε × l), where ε is molar extinction coefficient (M⁻¹cm⁻¹) and l is cuvette path length (cm).
What is the Pace method for estimating protein extinction coefficient? ▼
ε280 (M⁻¹cm⁻¹) = (5,500 × #Trp) + (1,490 × #Tyr) + (125 × #Cystine).