this is a spectrophotometry experiment from biochemsitry

Part A: Using Beer's law to Determine Concentration of an Unknown protein In this part of the experiment, you will use Beer's law to determine the concentration of a solution. 1) Adjust the spectrophotometer at 280nm, set the absorbance to zero using dH2O. 2) Obtain a solution of a protein with unknown concentration. 3) Measure the absorbance at 280nm. 4) Use the absorbance and molar extinction to determine the concentration (in mg/ml ) of the protein sample. (Hint: A=.c.b ) Extinction coefficient of BSA at 280nm is 43,824 M1cm1 Part B: Setting Up a Standard Curve In this part you will set up a standard curve for a protein determination, called the Biuret method. The protein standard will be bovine serum albumin, BSA, which is a generic protein that most people use for their protein assays. We usually set up a series of tubes with varying amounts of BSA and a constant amount of Biuret reagent. By plotting the number of milligrams or micrograms of BSA on the x-axis and the corrected absorbance on the y-axis, we can determine the concentration of unknown proteins from the graph. 1) Set up a protocol as in the following table: 2) Vortex immediately after adding the reagent to each tube. Do not wait until you have added it to all tubes. 3) Let the tubes sit about 10 min before reading the absorbencies. Once the color develops, it is stable for over an hour. 4) By using a suitable spectrophotometer fixed at wavelength 595nm, measure the absorbance of the sample and standards using a blank as a reference (auto zero). 5) Make a plot to determine how much BSA you can add without the curve straying from linear. You may find that it was linear through your highest standard (100uL), but it may also veer off at a lower volume of standard. If the absorbance of the unknown BSA is higher than your highest standard point that is on the straight part of the curve, you will need to make up another tube using less of the unknown BSA. You also want corrected absorbance of your unknown to be 0.8 or less. - What is the advantage and disadvantage of the method that you used to quantify the protein concentration ? compare this method with other methods for protein quantification