User:Mary Mendoza/Notebook/CHEM572 Exp. Biological Chemistry II/2013/04/17

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(Protocol)
(Objective)
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==Objective==
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* A run of 40 μM adenosine with 50 μM varying inhibitors were conducted using the UV 2500 Shimadzu spectrophotometer.
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* Three runs of each inhibitor were taken; the average of the runs were taken. The standard deviation were calculated and added as error bars.
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* Produce histogram
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==Protocol==
==Protocol==

Revision as of 18:40, 8 May 2013

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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.
  • Cuvette Mixture no Inhibitor

Image:CuvetteMixtureNoInhibitor.png

  • Cuvette Mixture with Inhibitor

Image:CuvetteSolnInhibitorHisto.png

  • Inhibitors Ran today:
  1. 50uM [2,3-bis(actyloxy)benzoic aicd] [PH003446]
  2. 50uM [Luteolin] [AMB18511398]
  3. 50uM [Morin] [AMB18511704]
  4. 50uM [Robinetin] [AMB18511408]
  5. 50uM [5-Hydroxy-2-methylbenzoic acid] [Aldrich 696366-1G]

Data


Notes

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



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