User:Dhea Patel/Notebook/CHEM 572: ADA&Inhibitor Kinetics/2013/04/09

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Objective

  • Using solutions made last week, a run of 40 μM adenosine with 25 μM varying inhibitors were conducted using the UV 2500 Shimadzu spectrophotometer.
  • Three runs of each inhibitor were taken; the average of the runs were taken. The standard deviation were calculated and added as error bars.
  • Produce histogram

Protocol

  • The reagents were added in the following sequence:
    • 1.78 mL of phosphate buffer
    • 600 μL of 200 μM adenosine (final concentration 50 μM)
  • This was allowed to mix through venting and placed on the chamber for absorbance reading.
    • 300 μL of phosphate buffer (or inhibitor dissolved in phosphate buffer)
    • 300 μL of dimethyl sulfoxide (DMSO) (or inhibitor dissolved in DMSO)
  • The cuvette contents were mixed by venting and placed on the chamber for absorbance reading.
    • 20 μL of adenosine deaminase (ADA) was added to catalyze the reaction
  • The kinetics of the reaction was taken using the UVProbe software.

Description

  • Cuvette Mixture with Inhibitor

Image:InhibitorCuvetteTable.png

  • Cuvette Mixture no Inhibitor

Image:CuvetteMixtureNoInhibitor.png

  • Inhibitors Ran today:
  1. Aspirin
  2. Kaempferol

Data

  • Image:409HistogramTable.png
  • Image:409AverageVelocityHisto.png
  • Image:409PercentChangeHisto.png

Notes

  • Solution Preparation/Calculations: Catherine Koenigsknecht
  • Cuvette preparation: Mary Mendoza
  • Data Evaluation: Dhea Patel
  • Overall Lab Assistance: Mike Nagle



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