User:Paul Rothenberg/Notebook/Pauls Notebook/2014/03/05

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Heme Extraction from Myoglobin

  • Preparation of Apoprotein:
  1. 24.3 mg of Tris was added to 4 mL of water to make a 0.05M solution to dissolve 200 mg of myoglobin
  2. 3.36 g of Sodium Bicarbonate was dissolved in 4L of H2O to make a 10 mM dialysis buffer
  3. The pH of the Tris solution was brought down to 2.3-2.5 at 0°C before 2-butanone extraction was performed
  4. 2-butanone was placed on ice for the extraction
  5. Acid/2-butanone extraction was performed on the protein solution with equal parts 2-butanone and protein
  6. The aqueous extract was then added to a dialysis cassette and placed in the dialysis buffer in the 0°C cold room and will sit overnight
    1. Approximately 3 mL of aqueous extract was extracted overall