Determinations of the Protein Concentration of a Cell Extract Using the 'Bradford Method'

ABSTRACTThe demand of the experiment is to harness out the concentration of vigorous phone extracts (Bovine Serum Albumin, BSA) utilize the Bradford order. Stock resolution of BSA was do by diluting the protein solution to pay off concentrations ranging from 0.0mg/cm3 to 1.0mg/cm3 (Peck, 1998). 5cm3 of the Bradford dye reagent was added to 100?l of the solution and allowed to stand for 28mins. The absorbance of each solution is measured at 595 nm with the following results: 0.00 mg/cm3 = 0.00, 0.2 mg/ cm3 = 0.21, 0.4 mg/ cm3 = 0.34, 0.6 mg/ cm3 = 0.56, 0.8 mg/ cm3 = 0.68, and 1.0 mg/ cm3 = 0.80. The value of the absorbance were plotted against protein concentrations and the resulting line was utilise to imagine the concentrations of the un cognise samples and the aim was achieved. INTRODUCTIONProtein concentration determination of samples is actually needed and it?s a great deal used i.e. to know the fargon of protein getable in a sample. Various methods for determining the protein concentrations in a sample ar available but variegate in ease of use, cost, and sensitivity. The close to commonly used methods are the Bradford balk, Lowry assay and the BCA assay. In this case, the Bradford method, which is a dye vertebral column method, is used.

It is faster, cheaper, and more sensitive, does non require heating, and has fewer mixing steps than the other methods above. This method is based on the binding of protein to a dye, leading to a shift in the absorbance fastness limit of the dye used. The dye, Coomassie blue, has a maximum of 464nm but when it binds to a protein, the maximum shifts to 595nm. After creating a example curve of protein concentrations of the known samples against absorbance, the concentrations of the unknown can be calculated. [Holme and Peck, 1998]MATERIALS... If you destiny to get a full essay, order it on our website: Orderessay

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