Bradford Assay Calculator
Our bio laboratory calculator computes bradford assay accurately. Enter measurements for results with formulas and error analysis.
Reviewed by Daniel Agrici, Founder & Lead Developer
Formula
[Protein] = (Abs_corrected - Intercept) / Slope x Dilution Factor
Where Abs_corrected = Sample Absorbance - Blank Absorbance, Slope and Intercept are derived from the BSA standard curve linear regression (Abs = Slope x [Protein] + Intercept), and Dilution Factor corrects for any sample dilutions made before measurement.
Worked Examples
Example 1: Standard Protein Quantification
Problem:A sample reads absorbance 0.45 at 595 nm with blank of 0.02. Standard curve: slope = 0.05 Abs/(ug/mL), intercept = 0.01. No dilution. Sample volume = 10 uL.
Solution:Corrected absorbance = 0.45 - 0.02 = 0.43\nConcentration = (0.43 - 0.01) / 0.05 = 8.40 ug/mL\nWith dilution factor 1: actual = 8.40 ug/mL\nTotal protein = 8.40 x (10/1000) = 0.084 ug
Result:Concentration: 8.40 ug/mL | Total protein: 0.084 ug
Example 2: Diluted Sample Analysis
Problem:A 1:5 diluted sample reads 0.62 absorbance with blank 0.03. Standard curve slope = 0.04, intercept = 0.02. Sample volume = 20 uL.
Solution:Corrected absorbance = 0.62 - 0.03 = 0.59\nConcentration (diluted) = (0.59 - 0.02) / 0.04 = 14.25 ug/mL\nActual concentration = 14.25 x 5 = 71.25 ug/mL\nTotal protein = 71.25 x (20/1000) = 1.425 ug
Result:Actual concentration: 71.25 ug/mL | Total protein: 1.425 ug
Frequently Asked Questions
How do I create a standard curve for the Bradford assay?
To create a standard curve, prepare serial dilutions of a known protein standard, typically bovine serum albumin (BSA), at concentrations ranging from 0 to 25 micrograms per milliliter for the micro assay or 0 to 2000 micrograms per milliliter for the standard assay. Add Bradford reagent to each dilution following the manufacturer protocol and incubate for five to ten minutes at room temperature. Measure absorbance at 595 nm against a blank containing only reagent and buffer. Plot absorbance on the y-axis versus concentration on the x-axis. Fit a linear regression line to determine the slope and y-intercept. The resulting equation allows you to calculate unknown protein concentrations from their absorbance values.
What are the limitations of the Bradford assay?
The Bradford assay has several notable limitations. First, the dye-protein response is not perfectly linear, especially at higher concentrations above 25 micrograms per milliliter in the micro assay. The assay shows protein-to-protein variation because dye binding depends on amino acid composition, so proteins with different proportions of arginine, lysine, and hydrophobic residues will produce different absorbance values at the same concentration. Detergents like SDS above 0.1 percent, strongly alkaline solutions, and some lipids can interfere with the assay. The standard curve should ideally use the same protein as the sample or a closely related one. Additionally, the color develops rapidly but is not perfectly stable over time.
References
Reviewed by Daniel Agrici, Founder & Lead Developer · Editorial policy