Cell Biology Lab [BIOL440-02 In-Class Exercise Time Expected to be allotted: 1:3

Question

Cell Biology Lab [BIOL440-02 In-Class Exercise Time Expected to be allotted: 1:30 -2 hr. Please read carefully and you can use your pc to use excel and generate the plot and the equation specified in this exercise You can work in groups of 2 and no more than 3; however, an individual report is expected from each member of the group. Total Protein Determination: Standard Calibration Curve Generation You are about to standardize the response of a spectrophotometer (auto-plate reader) to measure absorbance of total proteins in 6 samples. Using a primary bovine serum albumin (BSA) standard at a concentration of 2mg/ml A. Prepare 5 working standards of BSA at the following concentrations with a total volume of: 20 L: 1) 0.0 g/uL 2) 0.5 g/uL 3) 1.0 g/L 4) 1.5 g/L 5) 2.0 g/L B. Assuming that the absorbance (measured by the auto-plate reader) for each of the above standards is as follows: BSA Standard Concentration (ug/uL) ,Absorbance (Abs Units 0.0 0.5 ug/u 1.0 ug/u 1.5 ug/ul 0.0005 0.23 0.48 0.70 0.94 And knowing that the spectrophotometer (auto-plate reader) response in Absorbance units is proportional on BSA standard concentration: y-k) Where y is absorbance units and x is concentration of BSA standard: Prepare a plot and generate the best-fit line (calibration curve) and show the best-fit line equation (you can use excel to generate the plot (igure) and the Best-Fit Line Equation in the form of: y=mx + b Where m is the slope and b is the y-intercept

Explanation / Answer

Here we can find the volume V1 needed from stock of S1 concentration to prepare V2 volume of s2 concentration by using relation V1S1=V2S2

A) given stock (primary concentration of BSA standard solution) S1 =2mg/ml = 2000ug/ml = 2000ug/10000ul =2ug/ul

by using above said formula we can calculate how much volume will need to be pipette out form stock solution to prepare required volume (20ul) of required standard BSA solution which will come out as

1) for 20ul, 0.00ug/ul BSA solution, the volume of water will be needed = 20ul and volume will need to be pipette out form stock solution= 0ul

2) for 20ul, 0.5 ug/ul BSA solution, the volume of water will be needed = 15ul and volume will need to be pipette out form stock solution= 5ul

3)for 20ul, 1.00ug/ul BSA solution, the volume of water will be needed = 10ul and volume will need to be pipette out form stock solution =10ul

4)for 20ul, 1.5ug/ul BSA solution, the volume of water will be needed = 5ul and volume will need to be pipette out form stock solution=15ul

5)for 20ul, 2ug/ul BSA solution, the volume of water will be needed = 0ul and volume will need to pipette out form stock solution =20ul

B) when you will plot a graph from given data using excel you will get an equation y = 0.4698x + 0.0003

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