1. What Is Protein Concentration Calculation?

Protein concentration calculation using spectrophotometry is a fundamental technique in biochemistry and molecular biology laboratories. It allows researchers to quantify protein samples based on their absorbance of light at specific wavelengths.

2. How Does The Calculator Work?

The calculator uses the Beer-Lambert law formula:

Where:

• Conc — Protein concentration in milligrams per milliliter (mg/mL)
• Absorbance — Measured absorbance value (unitless)
• Extinction coefficient — Protein-specific constant (per mg/mL/cm)

Explanation: This formula calculates protein concentration based on the direct relationship between absorbance and concentration, using the protein's specific extinction coefficient.

3. Importance Of Protein Concentration Measurement

Details: Accurate protein quantification is essential for various applications including enzyme kinetics studies, protein purification, Western blotting, and ensuring consistent results in experimental procedures.

4. Using The Calculator

Tips: Enter the measured absorbance value and the protein-specific extinction coefficient. Both values must be positive numbers. Common extinction coefficients range from 0.5 to 2.0 per mg/mL/cm for most proteins.

5. Frequently Asked Questions (FAQ)

Q1: What is an extinction coefficient?
A: The extinction coefficient is a measure of how strongly a protein absorbs light at a specific wavelength. It is unique to each protein and depends on its amino acid composition.

Q2: At what wavelength should I measure absorbance?
A: For most protein concentration measurements using this method, absorbance is typically measured at 280 nm, which detects aromatic amino acids (tryptophan and tyrosine).

Q3: How do I find the extinction coefficient for my protein?
A: Extinction coefficients can be calculated from the protein's amino acid sequence using online tools, found in literature, or provided by protein databases.

Q4: What are the limitations of this method?
A: This method assumes the protein is pure and may be inaccurate if contaminants that absorb at 280 nm are present. It also requires knowledge of the exact extinction coefficient.

Q5: Are there alternative methods for protein quantification?
A: Yes, other methods include Bradford assay, BCA assay, Lowry assay, and quantitative amino acid analysis, each with different advantages and limitations.