Team:Caltech/Protocols/Measuring H2O2

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# 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.  
# 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.
#* NB: Dilute culture supernatants as appropriate to bring measurements into the assay’s linear range.
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[[Image:H2O2 standards example.png|thumb|center|400px|An example of a typical standard curve generated by the assay.]]
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Measuring H2O2


Note: Based on manufacturer's [http://www.piercenet.com/files/0526dh4.pdf instructions].

  1. Spin down approximately 100 uL of cell culture
  2. 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].
  3. After incubating the reaction at room temperature for at least 20 minutes, read on a plate reader (Spectramax M2) at 595nm.
  4. 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.
An example of a typical standard curve generated by the assay.
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