User:Matt Hartings/Notebook/Photosynthesis/2013/10/25

From OpenWetWare
Jump to: navigation, search
Hartings AU Photosynthesis Lab Header.png Protein Re-engineering Report.pngMain project page
Resultset previous.pngPrevious entry      Next entryResultset next.png



Objective

Take kinetic data for Mb (lyophilized from KCl/phosphate buffer) catalyzed o-phenylenediamine + H2O2 reaction in water (45C) and Mb (lyophilized from KCl/tris buffer) catalyzed o-phenylenediamine + H2O2 reaction in water (35, 45C).

Procedure and Data

sample 1

  1. Stock solutions:
    1. Myoglobin
      1. 1.4mg (Mb:KCl lyophilized from Phosphate, 5mg:500mg) in 1.0mL water
    2. OPD
      1. 2.3mg OPD in 2.3mL water
    3. H2O2
      1. 0.114mL 30% H2O2 in 0.887mL water
  2. Experimental Run
    1. Sample chamber set to 45C and to stir
    2. N2 run through the cuvette holder to reduce condensation on cuvette
    3. 2.025mL OPD stock solution
    4. 0.75mL H2O2 stock solution
      1. Start Data Collection
      2. 1ms Integration
      3. 10 scan average
      4. Wait 1 minute after each scan
      5. Stop after 2 hours
    5. Files saved as: Mb_OPD_H2O2_phosphate_water_45_SCANNUMBER.txt
    6. 0.225mL Mb stock solution added just before the third scan
    7. This is denoted in the data file for the third scan

sample 2

  1. Stock solutions:
    1. Myoglobin
      1. 1.2mg (Mb:KCl lyophilized from Tris, 5mg:500mg) in 1.0mL water
    2. OPD
      1. 3.0mg OPD in 3.0mL water
    3. H2O2
      1. 0.114mL 30% H2O2 in 0.887mL water
  2. Experimental Run
    1. Sample chamber set to 45C and to stir
    2. N2 run through the cuvette holder to reduce condensation on cuvette
    3. 2.025mL OPD stock solution
    4. 0.75mL H2O2 stock solution
      1. Start Data Collection
      2. 1ms Integration
      3. 10 scan average
      4. Wait 1 minute after each scan
      5. Stop after 2 hours
    5. Files saved as: Mb_OPD_H2O2_tris_water_45_SCANNUMBER.txt
    6. 0.225mL Mb stock solution added just before the third scan
    7. This is denoted in the data file for the third scan

sample 3

  1. Stock solutions:
    1. Myoglobin
      1. same as above
    2. OPD
      1. 2.3mg OPD in 2.3mL water
    3. H2O2
      1. 0.114mL 30% H2O2 in 0.887mL water
  2. Experimental Run
    1. Sample chamber set to 35C and to stir
    2. N2 run through the cuvette holder to reduce condensation on cuvette
    3. 2.025mL OPD stock solution
    4. 0.75mL H2O2 stock solution
      1. Start Data Collection
      2. 1ms Integration
      3. 10 scan average
      4. Wait 1 minute after each scan
      5. Stop after 2 hours
    5. Files saved as: Mb_OPD_H2O2_tris_water_35_SCANNUMBER.txt
    6. 0.225mL Mb stock solution added just before the third scan
    7. This is denoted in the data file for the third scan