XXXXXXXXXXTable 1: Standard and Unknown Protein Samples and Their Abso
ance
BI101 Latest 2014.pdf
57
DETERMINATION OF
PROTEIN CONCENTRATION
IN MILK PRODUCTS
OBJECTIVES
3. Cali
ate and use a spectrophotometer.
4. Plot a “standard” curve.
5. Use a standard curve to determine the protein concentration of several milk products.
INTRODUCTION
qualitative measurement of the presence of proteins.
quantitative
proteins bind to a dye called Coomassie Blue that is used in the Bradford protein assay. This
Determining protein concentration is of importance to biologist studying enzymes. Enzymes are
7
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Lab Section___________________________
58
Lab Exercise 7
RELEVANCE
The ability to precisely measure quantities of nutrients such as proteins is important in
quantitatively assess foods enabling us to tailor diets to the needs of patients, the elderly, and
your results to nutritional labels.
PROCEDURE
concentration of protein for each solution, a graph results. This is called a standard curve.
spectrophotometer.
Micropipette Practice:
dispensing volumes from 10-100
I. STANDARD CURVE AND SAMPLE PREPARATION
concentration.
1. Mix the tube of Coomassie Blue dye solution by inverting 3-4 times.
2. Next, use the micropipette of 1 mL capacity to add 3 mL of Coomassie Blue dye solution
to 9 spectrophotometer cuvettes labeled 0, 1, 2, 3, 4, 5, M1, M2, and M3.
L of each protein standard
0, 1, 2, 3, 4, and 5 using the XXXXXXXXXXL micropipette into the co
esponding cuvettes by
Be sure
to use a fresh tip for each sample.
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Protein Concentration in Milk
Table 1: Standard and Unknown Protein Samples and Their Abso
ance
Cuvette
Label
Type of
Protein
Sample
Coomassie
Blue Sample
Protein
Concentration
0 Standard 3 mL 60 L of tube 0
1 Standard 3 mL 60 L of tube 1
2 Satndard 3 mL 60 L of tube 2
3 Standard 3 mL 60 L of tube 3
4 Standard 3 mL 60 L of tube 4
5 Standard 3 mL 60 L of tube 5
M1 3 mL 60 L of tube M1
M2 3 mL 60 L of tube M2
M3 3 mL 60 L of tube M3
M1, M2 and M3 respectively by touching the tip of the pipette to the inside of the cuvette
Be sure to use a fresh tip for each sample.
5. vortex each tube so that the mixture produces a uniform color.
inaccurate data.
II. DETERMINING THE PROTEIN CONCENTRATION OF UNKNOWN
SAMPLES
1. Using abso
ance data from Table 1, create a standard curve by plotting the abso
ance
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Lab Exercise 7
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Protein concentration (mg/mL)
Table 2: Unknown Milk Protein Samples
Samples Values
Estimated Protein
M1
M2
M3
61
Protein Concentration in Milk
III. CALCULATIONS
Table 3: True Concentrations of Unknown Samples
Sample
Estimate of Protein
Concentration from Multiplied by the Dilution Factor Equals
Final Concentration
M1 × 50
M2 × 50
M3 × 50
actual composition of the samples if presented nutritional labels from several milk products.
Table 4: Converted Concentrations of Unknown Samples
Sample Final Concentration Multiplied by the Conversion Factor Equals
Concentration
M1 ×
M2 ×
M3 ×
7 g per serving. This value is directly comparable to the provided nutritional labels. Using this
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Lab Exercise 7
Table 5: Serving-Size Concentrations for the Unknown Samples
Sample Concentration Times Number of mL per serving Equals
Concentration
M1 ×
M2 ×
M3 ×
actual identities to the class.
Table 6: Probable Identities of Unknown Samples
sample
Calculated class average for the Probable milk item Were you
M1
M2
M3
IV. QUESTIONS
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Lab Section: