User:Megan L. Channell/Notebook/Horseradish/2013/10/09

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Current revision (15:36, 10 October 2013) (view source)
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==Notes==
==Notes==
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This area is for any observations or conclusions that you would like to note.
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'''EHNA stock'''
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# 5mg EHNA in 1mL DMSO ---> 15.9mM EHNA
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# (1.9μL)(15.9mM EHNA)=(10mL)(C2). C2 ---> 3μM EHNA
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The reaction samples will contain roughly 1nM EHNA.
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Use categories like tags. Change the "Course" category to the one corresponding to your course. The "Miscellaneous" tag can be used for particular experiments, as instructed by your professor. Please be sure to change or delete this tag as required so that the categories remain well organized.
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[[Category:CHEM571]]
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[[Category:Adenosine]]
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[[Category:Inosine]]
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Current revision

Biomaterials Design Lab Main project page
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Objective

Continue work from yesterday, but to observe the ADA kinetics with an inhibitor.

Procedure

Creating the Adenosine Stock

  1. Add 0.1073 g of adenosine to a 10 ml volumetric flask. Fill with 50 mM phosphate buffer at pH 7.4. This created a stock solution of 0.04 M.
  2. Add 10.00 μL of the 0.04 M adenosine stock to a 10 mL volumetric flask. Fill with 50 mM phosphate buffer at pH 7.4. This created a final solution of 40 μM.

Enzyme Kinetics Measurement

  1. Add 3 mL of 40 μM adenosine solution to the cuvette and 30 μL of inosine.
  2. Start kinetics measurement:
  • 1 ms integration
  • 10 scan average
  • Set "Save the first available scan every" to 15 seconds
  • Set "Stop after this amount of time" to 10 minutes
  • Set "File Type" to Tab Delimited
  • Just before 1 minute, add 1.5 μL of 1 nM EHNA.

Data

Image:Cmj_10.09.13_ADAwithinhibitor.png

Notes

EHNA stock

  1. 5mg EHNA in 1mL DMSO ---> 15.9mM EHNA
  2. (1.9μL)(15.9mM EHNA)=(10mL)(C2). C2 ---> 3μM EHNA

The reaction samples will contain roughly 1nM EHNA.




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