Protein Concentration Calculator
Enter the 280 nm absorbance, molar extinction coefficient, molecular weight, dilution factor, and path length to compute protein concentration (mg/mL and molar) via the Beer–Lambert law, with a reference table of common proteins.
Input Data
Results
At a glance:Protein concentration can be quickly estimated from UV absorbance: aromatic amino acids (tryptophan, tyrosine) absorb specifically at 280 nm. By the Beer–Lambert law A = ε·c·b, first solve molar concentration c = A / (ε·b), then multiply by molecular weight MW to convert to mass concentration (mg/mL), and multiply by dilution factor n to recover the stock concentration: C = A / (ε·b) × MW × n.
Formula
Molar concentration: c = A ÷ (ε × b) (mol/L).
Mass concentration: C = c × MW × n (mg/mL, since g/L = mg/mL).
Combined: C = A ÷ (ε × b) × MW × n.
$$A = \varepsilon \, c \, b$$$$C = \dfrac{A}{\varepsilon \, b} \times \mathrm{MW} \times n$$How to Use
- Enter the protein absorbance A at 280 nm.
- Enter the molar extinction coefficient ε and molecular weight MW (look up or estimate from sequence), plus dilution factor and path length.
- The right panel shows mass concentration (mg/mL) and molar concentration (mol/L) instantly.
Reference: molar extinction coefficients and molecular weights of common proteins
| Protein | ε₂₈₀ (M⁻¹cm⁻¹) | Molecular weight (g/mol) |
|---|---|---|
| Bovine serum albumin (BSA) | 43824 | 66463 |
| Immunoglobulin IgG | 210000 | 150000 |
| Lysozyme | 38940 | 14313 |
| Cytochrome c | source-dependent | 12384 |
Extinction coefficients vary slightly with folding state and solvent; precise quantification is best cross-validated by BCA or Bradford assays.
Case Studies
Quantifying purified BSA
After purifying BSA, A280 = 0.5, path length 1 cm, undiluted (n=1). Use ε=43824, MW=66463.
c = 0.5 ÷ (43824 × 1) = 1.141×10⁻⁵ mol/L; C = 1.141×10⁻⁵ × 66463 ≈ 0.758 mg/mL.
This concentration can be used for downstream volume conversion or to judge whether concentration is needed.
High-concentration IgG antibody sample
Antibody stock absorbance too high; dilute 2× then measure A = 1.4 (ε=210000, MW=150000 for example).
Restore stock: C = 1.4 ÷ (210000 × 1) × 150000 × 2 = 2.0 mg/mL.
Always record and enter the dilution factor n, otherwise concentration is underestimated by half.
FAQ
Why does protein absorb at 280 nm?
Because the side chains of aromatic amino acids (tryptophan, tyrosine) absorb UV at 280 nm; the more such residues, the larger the extinction coefficient ε. Proteins lacking aromatic residues are unsuitable for this method.
Where do I get the molar extinction coefficient ε?
Estimate it from the amino-acid sequence using tools like ExPASy ProtParam (based on Trp, Tyr, and cystine counts), or look up literature values. The more accurate ε, the more reliable the concentration estimate.
How do mg/mL and molar concentration convert?
Mass concentration (mg/mL = g/L) = molar concentration (mol/L) × molecular weight (g/mol). This calculator outputs both for easy cross-checking.
Which is more accurate, UV or Bradford/BCA?
UV (A280) is fast and non-consuming but needs a known ε; Bradford/BCA are colorimetric, need a standard curve but are more general for unknown proteins. For precise quantification, cross-validate the two methods.
Absorbance too high (>1.5), what to do?
Above the linear range it underestimates concentration; dilute the sample, re-measure, and enter the actual factor in 'dilution factor n' — the tool auto-restores the stock concentration.
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References
Content review: Calculatorism Science Team. Results are for reference only; please refer to the relevant authorities for the official figures.