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| - | Approximately 100 uL of cell culture was spun down. A colorimetric assay was performed using the PeroXOquant™ Quantitative Peroxide Assay Kit (Pierce) as per the manufacture’s instructions in a 96 well plate. After incubating the reaction at room temperature for at least 20 minutes, it was read on a plate reader (Spectramax M2) at 595nm. The concentration of hydrogen peroxide was calculated by reference to a standard (100uM, 33.3uM, 11.1 uM, 3.7 uM and 0.0 uM H2O2) made in the corresponding media. Culture supernatants were diluted as appropriate to bring measurements into the assay’s linear range.
| + | Note: Based on manufacturer's [http://www.piercenet.com/files/0526dh4.pdf instructions]. |
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| | + | # Spin down approximately 100 uL of cell culture |
| | + | # Measure peroxide concentration using a [http://www.piercenet.com/Products/Browse.cfm?fldID=02020301 colorimetric assay] and following the manufacturer's [http://www.piercenet.com/files/0526dh4.pdf instructions]. |
| | + | # After incubating the reaction at room temperature for at least 20 minutes, read on a plate reader (Spectramax M2) at 595nm. |
| | + | # Calculate the concentration of hydrogen peroxide by reference to a standard curve (100uM, 33.3uM, 11.1 uM, 3.7 uM and 0.0 uM H2O2) made in the corresponding media. |
| | + | #* NB: Dilute culture supernatants as appropriate to bring measurements into the assay’s linear range. |
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Revision as of 22:10, 17 October 2008
iGEM 2008
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Measuring H2O2
Note: Based on manufacturer's [http://www.piercenet.com/files/0526dh4.pdf instructions].
- Spin down approximately 100 uL of cell culture
- Measure peroxide concentration using a [http://www.piercenet.com/Products/Browse.cfm?fldID=02020301 colorimetric assay] and following the manufacturer's [http://www.piercenet.com/files/0526dh4.pdf instructions].
- After incubating the reaction at room temperature for at least 20 minutes, read on a plate reader (Spectramax M2) at 595nm.
- Calculate the concentration of hydrogen peroxide by reference to a standard curve (100uM, 33.3uM, 11.1 uM, 3.7 uM and 0.0 uM H2O2) made in the corresponding media.
- NB: Dilute culture supernatants as appropriate to bring measurements into the assay’s linear range.
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