Difference between revisions of "Peroxidase UV Vis assay"
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introduction...We are interested in measuring the peroxiase activity of a solution. We are going to isolate ovop from sea urchins and we need to know how actie the enzyme is. We are starting with HRP and ... | introduction...We are interested in measuring the peroxiase activity of a solution. We are going to isolate ovop from sea urchins and we need to know how actie the enzyme is. We are starting with HRP and ... | ||
==Standard Mixing Procedure== | ==Standard Mixing Procedure== | ||
| + | The mixture of Peroxidase UV-Vis assays requires the use of a Guaiacol stock solution and a hydrogen peroxide stock solution. The stock solutions are prepared as follows: | ||
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| + | Guaiacol Stock Solution | ||
| + | |||
| + | 49.5 microliters of Guaiacol is added to a 25 mL pH 8.0 buffer solution, which is comprised of 25 mL of DI water and a 8.0 pH buffer tablet that has been mixed thoroughly. | ||
| + | |||
| + | Hydrogen Peroxide Stock Solution | ||
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| + | 17 microliters of hydrogen peroxide is added to 1 mL of DI water. | ||
| + | |||
| + | Assay | ||
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| + | 5.0 mL Guaiacol stock solution is added to a vial. | ||
| + | |||
| + | 10 microliters of the hydrogen peroxide stock solution is added to the Guaiacol stock solution. | ||
| + | |||
| + | The desired amount of peroxidase is then added to the assay before the UV-Vis is ran. | ||
==Example Data== | ==Example Data== | ||
Revision as of 01:39, 12 November 2018
introduction...We are interested in measuring the peroxiase activity of a solution. We are going to isolate ovop from sea urchins and we need to know how actie the enzyme is. We are starting with HRP and ...
Standard Mixing Procedure
The mixture of Peroxidase UV-Vis assays requires the use of a Guaiacol stock solution and a hydrogen peroxide stock solution. The stock solutions are prepared as follows:
Guaiacol Stock Solution
49.5 microliters of Guaiacol is added to a 25 mL pH 8.0 buffer solution, which is comprised of 25 mL of DI water and a 8.0 pH buffer tablet that has been mixed thoroughly.
Hydrogen Peroxide Stock Solution
17 microliters of hydrogen peroxide is added to 1 mL of DI water.
Assay
5.0 mL Guaiacol stock solution is added to a vial.
10 microliters of the hydrogen peroxide stock solution is added to the Guaiacol stock solution.
The desired amount of peroxidase is then added to the assay before the UV-Vis is ran.