User:Carly M. Montanero/Notebook/CHEM-571/2013/10/08

Biomaterials Design Lab: Fall 2013

Main project page Previous entry      Next entry

Objective To observe and measure ADA turnover kinetics in the absence of an inhibitor. This work will be the basis of our comparison to ADA-AuNP turnover studies. From Dr. Hartings. Procedure Creating the Adenosine Stock Add 0.1071 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. Add 9.98 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 Add 3 mL of 40 μM adenosine solution to the cuvette. 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 30 μL of 0.01 units/mL ADA. Figures Notes Adenosine deaminase (ADA) stock 1.1mg (24.0 units in 1 mg) in 25mL of buffer --> 1.1 units/mL ADA ADA for experiments 100uL of stock ADA and 900uL buffer --> 0.11 units/mL ADA