“Compare the advantages and disadvantages of the Bradford dye-binding protein as
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“Compare the advantages and disadvantages of the Bradford dye-binding protein assay and the UV absorption assay.” For my Biochemistry lab report I need to compare these two spectroscopic methods, listing both their advantages and disadvantages “Compare the advantages and disadvantages of the Bradford dye-binding protein assay and the UV absorption assay.” For my Biochemistry lab report I need to compare these two spectroscopic methods, listing both their advantages and disadvantages For my Biochemistry lab report I need to compare these two spectroscopic methods, listing both their advantages and disadvantagesExplanation / Answer
Bradford dye-binding assay
advantages
1. Many protein solution shows highest peak at 280nm which require spectrophotometers which can measure the uv range and some of the will show light peak at 280 nm. To avoid all of the complications the protein samples are mixed with Bradford reagent and can measure the absorbance at visible range (580 nm)
2. The Bradford dye-binding process is simple and easy.
3. The dye binds with the protein to form stable interections.
4. It is low cost
5. It is highly sensitive to visible spectrophotometer
6. The experiment is took less time and can be done at room temperature.
disadvantages
1. concentration range is low 0 microgram to 2000 microgram.
2.The process can be inhibited by the detergents and basic medium.
3.Bradford assay dependable on protein sequence, if the protein does not have ideal number of aromatic residue it will not bind well to the dye.
4. The Bradford reagent stains the test tubes and cuvettes which is hard to remove and can interfere in the further experiments.
UV absorption assay
advantages
1. Aromatic amino acids , tryptophan, tyrosine, shows peak at this range (280 nm) .
2. It is quick and easy
3. No chemical reaction is required.
4. The proteins of different ratio of aromatic residues are more suited in this method contrary to Bradford dye-binding assay.
disadvantages
1. The method has medium sensitivity
2. It is disturbed by DNA molecules as nucleic acids have an adsorption maxima of 260 nm and at 280 nm their adsorption is also significant.
3. The wavelength range is very poor,some of the proteins have higher adsorption maxima in those cases this method not much suitable.
4. This method is highly sensitive to pH and ionic strength
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