2. BSA was used as a standard in the Bradford colorimetric assay. For comparison,
use the extinction
coefficient at 280 nm for BSA to calculate the concentration of
hemoglobin and lysozyme from their absorbance values at 280 nm (A280).
Compare these values to the actual concentrations of hemoglobin and lysozyme.
What do you conclude from this comparison?
3. Assumed that you have isolated the protein chymotrypsinogen from a cell lysate.
Discuss the advantages and disadvantages of the following protein assays.
a. Absorbance at 215 nm
b. Absorbance at 280 nm
c. The Bradford assay
4. Compare extinction coefficients for DNA (1 mg/ml, 260 nm) and the proteins (1
mg/ml, 280 nm). Explain any difference between the DNA and protein extinction
coefficients in the region of the UV spectrum.
5. Suppose you had a sample that contained equal concentrations of DNA and
protein. Sketch the absorption spectrum you would expect to see in the region
from 240 to 300 nm.
6. BSA has the following profile of amino acid composition compared with the
average of known vertebrate proteins:
AA BSA Average protein
Phenylalanine (Phe) 4.6% 4.0%
Histidine (His) 2.7% 2.9%
Lysine (Lys) 10.1% 7.2%
Arginine (Arg) 3.9% 4.2%
Tryptophan (Trp) 0.3% 1.3%
Tyrosine (Tyr) 3.4% 3.3%
Based upon the given table, is the BSA a good standard to use in the Bradford assay? Explain your answer.