Exp_3_Key - MCB120L Experiment 3 Write-up Key Part A: (i)...

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MCB120L Morand Experiment 3 Write-up Key Part A: (i) Properly constructed and properly labeled graph of: IU (y-axis) vs pH (x-axis). (ii) Given the name of the enzyme, Alkaline Phosphatase, a pH optimum above pH 8 is expected. Also, enzyme activity graphs examining optimum parameters such as temperature dependence, pH dependence, ionic strength dependence, etc, are typically “bell shaped” ( ie not linear!), that is, a sigmoid curve. Part B: 1. Determining the absorption coefficient, ε l , for p-nitrophenol on the microplates from class data. slope = ε l = ( Abs / μM) = ( 0.0126 μM -1 l -1 x l ) OR 0.0126 μM -1 (has typically ranged from 0.0125 μM -1 to 0.0131 μM -1 depending on data, data elimination & differences in the calculations performed by the Excel program) p-nitrophenol standard y = 0.0126x R 2 = 1 0.000 0.200 0.400 0.600 0.800 1.000 1.200 1.400 1.600 1.800 0.000 20.000 40.000 60.000 80.000 100.000 120.000 140.000 uM p-nitrophenol Abs 405 nm 2. (i) Calculating IU = (Abs 405 corrected / 0.0126 μM -1 )(250 x 10 -6 L) Calculating mg of alkaline phosphatase; 0.125 μg/μl stock concentration 1/1-dilution in rx: (10 μl)(0.125 μg/μl) = 1.25 μg = 1.25 x 10 -3 mg 1/2-dilution (1/2)(1.25 x 10
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Exp_3_Key - MCB120L Experiment 3 Write-up Key Part A: (i)...

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