Sod activity kit

Superoxide dismutase (SOD) is an endogenous antioxidant which scavenges the superoxide anion into hydrogen peroxide and oxygen. SOD activity was measured by means of a Biovision SOD activity kit (Mountain View, CA, USA), according to manufacturer’s instructions. Briefly, H9c2 cardiomyoblasts were lysed by the addition of 100 µL of 0.1 M trizma/hydrochloride (Tris/HCl) and 0.1 mg/mL phenylmethylsulfonyl fluoride (PMSF). Lysed cells were then centrifuged at 15,000× g for 5 min at 4 °C. Twenty microliter of the cell lysates were then transferred to a fresh 96-well plate with the addition of 200 µL WST and 20 µL Enzyme Working Solution (supplied in the kit), where after the cells were incubated at 37 °C for 20 min prior to measuring of SOD activity. Absorbance was read at an excitation and emission wavelength of 485 ± 20 nm/528 ± 20 nm with a Biotek ELX 800 plate reader using Gen 5 software.

Cells were harvested 48 hr post treatment of different concentration of 2-methoxyestradiol and equal concentrations of protein were used for SOD activity analysis, using SOD activity kit as per manufacturer's instructions (Biovision).