User:Klare Lazor/Notebook/Chem-496-001/2011/09/14
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ObjectiveTo determine the concentration of an unknown protein, a Bradford Assay was used. DescriptionTo determine the concentration of an unknown protein, a Bradford Assay was performed. Materials needed included Bradford Reagent, bovine serum albumen (BSA), and protein (MBP). Standard solutions of BSA (1mL each) in water of 10, 8, 6, 4, 2, and 1ug/mL were prepared. First, 3mL of a 10ug/mL stock solution was created. From this stock solution the standards, as well as the sample of unknown protein, were made. The sample of the unknown protein should be 1000 times too concentrated. Furthermore, a blank for the Bradford Reagent and the protein were prepared. A UV-visible spectra was taken of each solution with a wavelength range of 200-800nm. DataChart 1: Is a graph of Absorbance vs. Concentration at 595nm. From this we can use the line of best fit to determine the concentration of protein in the unknown. We eliminated one data point because it was irrelevant. Chart 2: Is a graph of Absorbance vs. Wavelength. Chart 3: Is a graph of molar absorbtivity vs. wavelength. This was found using the equation A=Ebc. Chart 4: Is a graph of Protein Absorbance in water vs. Wavelength. We can use the Absorbance of protein at 595nm to determine the molar absorbtivity, and eventually, find the concentration of protein in the unknown. Calculations: Concentration of Unknown
NotesObservations:
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