Enzyme protocol

Enzyme protocol - Enzyme Lab Protocol October 22, 2008...

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Enzyme Lab Protocol October 22, 2008 Biology 2112 Part A Objective : We are finding the absorbency of the different concentrations of p-nitrophenol and  make a    standard curve. Procedure : 1. Label six tubes  “1,2,3,4,5, Blank” 2. Prepare the dilutions of P-ntirophenol using the .05mM standard. The molarities should  be .0025mM, .005mM, .01mM and .05mM and all should have total volumes of 3mL. 3. Blank the spectrophotometer at 405nm with the buffer blank 4. Record the absorbencies of each dilution at 405nm  5. Make the standard curve Calculations : Data :
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Dilution Concentration (mM) Absorbance (@ 405nm) .0025 .136 .005 .211 .01 .351 .025 .830 .05 1.95 Part B Objective : We are to find the rate of reaction of p-nitrophenyl phosphate, by adding an enzyme  and buffer to follow the production rate of the substrate Procedure: 1. Prepare the solutions with the following dimensions and label accordingly: Sample 1  (without enzyme)- 0.8 mL buffer and 2mL substrate, Sample 1 blank – 2.8 mL buffer and  0.2 mL water, Sample 2 (with enzyme) – 0.8 mL buffer and 2 mL substrate, Sample 2  blank – 2.8 mL buffer and 0.2 mL serum. Mix properly. 2. Blank the instrument at 405nm. Add 0.2mL of water to sample 1, mix and record the  reading at time 0. Continue to read sample 1 at 1 minute intervals over a course of 10  minutes. Record results and remember to blank the instrument several times. Did the 
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This note was uploaded on 04/12/2009 for the course BIOLOGY 1111 taught by Professor Tanaka during the Fall '09 term at Temple.

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Enzyme protocol - Enzyme Lab Protocol October 22, 2008...

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