1. What is the Nanodrop Absorbance Method?

The Nanodrop absorbance method is a quick and convenient technique for measuring protein concentration using ultraviolet absorbance at 280 nm. This method leverages the fact that proteins containing aromatic amino acids (tryptophan, tyrosine) absorb light at this wavelength.

2. How Does the Calculator Work?

The calculator uses the simple absorbance formula:

Where:

• \( A_{280} \) — Absorbance reading at 280 nm from Nanodrop spectrophotometer
• 1.0 — Extinction coefficient (assuming 1 mg/mL solution gives A280 = 1.0)

Explanation: This calculation assumes that a 1 mg/mL protein solution has an absorbance of 1.0 at 280 nm, which is a common approximation for many proteins.

3. Importance of Protein Concentration Measurement

Details: Accurate protein concentration measurement is essential for various biological and biochemical applications, including protein purification, enzyme kinetics studies, Western blotting, and protein quantification for experimental setups.

4. Using the Calculator

Tips: Enter the A280 absorbance reading from your Nanodrop spectrophotometer. Ensure the reading is taken at 280 nm and the sample is properly blanked with appropriate buffer.

5. Frequently Asked Questions (FAQ)

Q1: Why use A280 for protein concentration measurement?
A: A280 measurement is quick, requires small sample volumes, and is non-destructive, allowing sample recovery for further experiments.

Q2: What is the accuracy of this method?
A: The accuracy depends on the protein's specific extinction coefficient. The 1.0 value is an approximation - for precise measurements, use the protein's actual extinction coefficient.

Q3: When is this method not appropriate?
A: Not suitable for proteins lacking aromatic amino acids, or when the sample contains other UV-absorbing contaminants (nucleic acids, certain buffers).

Q4: How does this compare to other protein assays?
A: Faster than Bradford or BCA assays but less specific. Ideal for pure protein samples without interfering substances.

Q5: What sample volume is needed for Nanodrop measurement?
A: Typically 1-2 μL, making it ideal for precious samples where volume conservation is important.