1. What is the A280 Protein Concentration Method?

The A280 method estimates protein concentration by measuring absorbance at 280nm, where aromatic amino acids (tryptophan and tyrosine) absorb UV light. This method is quick and non-destructive, making it ideal for blood protein analysis.

2. How Does the Calculator Work?

The calculator uses the Beer-Lambert law:

Where:

• \( C \) — Protein concentration (mg/ml)
• \( A_{280} \) — Absorbance at 280nm (dimensionless)
• \( \epsilon \) — Extinction coefficient (ml mg^-1 cm^-1)
• \( l \) — Path length (cm)

Explanation: The absorbance is directly proportional to the protein concentration when the extinction coefficient and path length are known.

3. Importance of Protein Quantification

Details: Accurate protein concentration measurement is essential for biochemical experiments, clinical diagnostics, and quality control in biopharmaceutical production.

4. Using the Calculator

Tips: Enter absorbance at 280nm, extinction coefficient (1.0 ml mg^-1 cm^-1 is typical for many blood proteins), and path length (usually 1.0 cm for standard cuvettes). All values must be positive.

5. Frequently Asked Questions (FAQ)

Q1: What is a typical extinction coefficient for blood proteins?
A: For serum albumin, ε ≈ 0.66 ml mg^-1 cm^-1. For antibody solutions, ε ≈ 1.4 ml mg^-1 cm^-1.

Q2: Why is the path length important?
A: Longer path lengths result in greater absorbance. The standard is 1 cm, but microvolume instruments may use shorter paths.

Q3: What are limitations of the A280 method?
A: Nucleic acid contamination can interfere, and different proteins have different extinction coefficients based on their aromatic amino acid content.

Q4: How accurate is this method?
A: With a known extinction coefficient, accuracy is typically ±10%. For unknown mixtures, Bradford or BCA assays may be better.

Q5: Should samples be diluted?
A: Ideal absorbance values are between 0.1-1.0. Samples outside this range should be diluted or concentrated accordingly.