User:Michael F. Nagle/Notebook/Chem 571/2012/10/02: Difference between revisions

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==Procedure==
==Procedure==
#HRP procedure<sup>1</sup> was followed. Time was plotted against absorbance on the UV/Vis spectroscoper.
#HRP procedure<sup>1</sup> was followed. Time was plotted against absorbance on the UV/Vis spectroscoper.
#1mL horseradish peroxidase was dissolved in 1mL distilled water.  
#1mg horseradish peroxidase was dissolved in 1mL distilled water.  
#A .2M buffer of sodium phosphate was made at a pH of 7.
#A .2M buffer of sodium phosphate was made at a pH of 7.
#.0017M hydrogen peroxide solution was made by adding 1mL 30% hydrogen peroxide to 100mL distilled water. 1mL of hydrogen peroxide solution was diluted to 50mL with 49mL of sodium phosphate buffer for 34μM solution.  
#.0017M hydrogen peroxide solution was made by adding 1mL 30% hydrogen peroxide to 100mL distilled water. 1mL of hydrogen peroxide solution was diluted to 50mL with 49mL of sodium phosphate buffer for 34μM solution.  

Revision as of 22:10, 6 December 2012

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Objective

  • Prepare reagents for horseradish peroxidase (HRP) assay
    • Complete HRP assay with and without gold nanoparticles to see if they interact with HRP's active site

Procedure

  1. HRP procedure1 was followed. Time was plotted against absorbance on the UV/Vis spectroscoper.
  2. 1mg horseradish peroxidase was dissolved in 1mL distilled water.
  3. A .2M buffer of sodium phosphate was made at a pH of 7.
  4. .0017M hydrogen peroxide solution was made by adding 1mL 30% hydrogen peroxide to 100mL distilled water. 1mL of hydrogen peroxide solution was diluted to 50mL with 49mL of sodium phosphate buffer for 34μM solution.
  5. 698μM iodophenol solution was made in 1mL DMSO, since it's not soluble in water. 2.5mM 4-aminoantipyrine solution was prepared with the sodium phosphate buffer.

Discussion

  1. The HRP assays were not completed today, but will be 10/02, with the reagents prepared today

References

1. Horseradish Peroxidase Assay. Gold Biotechnology File:3760-Protocol HRP AbsorbanceAssay.pdf


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