A 20 mL protein fraction recovered from a column chromatography was analyzed for protein usingthe BCA method. The following data were the means of a duplicate analysis using BSA as astandard:2The average absorbance of a 1 mL sample was 0.44. Calculate protein concentration (mg/mL) andtotal protein quantity of this column fraction. (Hint: use Excel to establish the calibration curve)

To calculate the protein concentration (mg/mL) and total protein quantity of the column fraction, we need to establish a calibration curve using the BCA method and the provided data.

1. Start by plotting a calibration curve using Excel. The x-axis should represent the protein concentration in mg/mL, and the y-axis should represent the average absorbance values.

2. Use the duplicate analysis data for BSA as a standard to plot the calibration curve. Since the average absorbance of a 1 mL sample of BSA was 0.44, you can plot this point on the graph.

3. Repeat step 2 for different known concentrations of BSA, such as 0.1 mg/mL, 0.2 mg/mL, 0.3 mg/mL, and so on. Measure the average absorbance for each concentration and plot these points on the graph.

4. Once you have plotted all the points, add a trendline to the graph. This trendline will help you determine the equation of the line, which relates the protein concentration to the absorbance values.

5. Now, using the equation of the trendline, you can calculate the protein concentration of the column fraction. Plug in the average absorbance value of the 1 mL sample (0.44) into the equation and solve for the protein concentration in mg/mL.

6. To calculate the total protein quantity of the column fraction, multiply the protein concentration (mg/mL) by the volume of the fraction (20 mL). This will give you the total protein quantity in mg.

By following these steps and using Excel to establish the calibration curve, you can calculate the protein concentration and total protein quantity of the column fraction.