Need help with prelab calculations for biochemistry Polystyrene curettes Protein

ID: 532432 • Letter: N

Question

Need help with prelab calculations for biochemistry Polystyrene curettes Protein Standards (BSA -bovine serum albumin) Bradford reagent Egg whites or milk or other liquid protein unknown PBS (phosphate buffered saline) buffer The unknown samples probably have too much protein and are out of the range of Beers Law. These samples must be diluted, but by how much? Since the concentration is unknown, a dilution series will be assayed so that at least one sample is within the linear range of the standard curve. generally, you might want to prepare a sample that is 2x too high or 2x too low just in case. For this lab, dilute your samples 1: 10, 1: 100 and 1: 1000. Be sure to label your tubes appropriately! Place 20 mu l of each diluted sample into clean, labeled tubes. Using the chart below, you will prepare a series of samples of protein of known concentrations referred to as "standard proteins". Pipet 20 mu l of each into a labeled tube. Calculate how much of each protein you added to each tube. Add i ml Bradford Reagent to all tubes, 1-8 and the 20 mu l of each diluted unknowns. Mix the colorometric reaction should be complete after about 5 minutes, and the color should remain stable for about 1 hour. After turning on the spectrophotometer allow the machine to warn up for at least 5 minutes before making any measurements. Set the Wavelength: set the Wavelength using the nm and buttons.

Explanation / Answer

We know that 1 µL = 10-6 L = 10-3 mL.

We are to find out the amount (in milligrams) of the added standard protein in each of the prepared solutions.

20 µL of PBS buffer was added to test tube #1; hence the amount of added protein = 0.

Take test tubes #2 and #3 as examples.

Amount of protein added = (concentration of protein in mg/mL)*(volume of standard protein added to each tube).

Take test tube#2.

Amount of protein = (0.125 mg/mL)*(20 µL)*(1 mL/1000 µL) = 0.0025 mg.

Take test tube#3.

Amount of protein = (0.250 mg/mL)*(20 µL)*(1 mL/1000 µL) = 0.0050 mg.

Fill in the table as below:

Test tube #

Volume of standard protein added (µL)

Concentration of standard protein added (mg/mL)

Amount of standard protein added in millimgrams = (concentration in mg/mL)*(volume in mL)

PBS

0.0000

0.125

0.0025

0.250

0.0050

0.500

0.0100

0.750

0.0150

1.000

0.0200

1.500

0.0300

2.000

0.0400