I am presently working with H9C2 cell. Now, I want to quantify superoxide dismutase enzyme after exposure of hypoxic condition to H9C2 cells. So, which methods should I have to use?
Measurement of superoxide dismutase, catalase and glutathione peroxidase in cultured cells and tissue( Nature Protocols 5, 51–66 (2010)doi:10.1038/nprot.2009.197)
Abstract: Cells contain a large number of antioxidants to prevent or repair the damage caused by reactive oxygen species, as well as to regulate redox-sensitive signaling pathways. General protocols are described to measure the antioxidant enzyme activity of superoxide dismutase (SOD), catalase and glutathione peroxidase. The SODs convert superoxide radical into hydrogen peroxide and molecular oxygen, whereas the catalase and peroxidases convert hydrogen peroxide into water. In this way, two toxic species, superoxide radical and hydrogen peroxide, are converted to the harmless product water. Western blots, activity gels and activity assays are various methods used to determine protein and activity in both cells and tissue depending on the amount of protein required for each assay. Other techniques including immunohistochemistry and immunogold can further evaluate the levels of the various antioxidant enzymes in tissues and cells. In general, these assays require 24–48 h to complete.
A new direct method for determining superoxide dismutase activity by measuring hydrogen peroxide formation( https://doi.org/10.1016/0009-2797(93)90112-CGet rights and content)
Abstract: A new, direct method for determining superoxide dismutase activity is presented in this study. This method is based on measurement of one of the products of the superoxide dismutase reaction, hydrogen peroxide. Hydrogen peroxide is quantitated using a coupled reaction where horseradish peroxidase catalyzes the formation of a fluorescent product, 6,6′-diOH-(1,1′-biphenyl)-3,3′-diacetic acid, from 4-OH-phenylacetic acid and hydrogen peroxide. Substrate for superoxide dismutase is provided by reduction of oxygen during the autoxidation of riboflavin in the presence of UV light. A linear correlation between the amount of superoxide dismutase (200 ng – 6 μg) and of hydrogen peroxide was found with this method
Superoxide Dismutase Activity Assay Kit (Colorimetric ... - Abcam www.abcam.com › … › Cellular metabolic process Superoxide Dismutase Activity Assay Kit ... the inhibition activity of SOD can be determined by a colorimetric method. ... Superoxide dismutase measured in biofluids
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