How do you do SOD assay?
SOD activity can be measured by both activity assays and activity gels. In the biochemical method, xanthine-xanthine oxidase is used to generate O2•− and nitroblue tetrazolium (NBT) reduction is used as an indicator of O2•− production.
How do you test for superoxide dismutase?
SOD activity was detected by incubating the gel in staining buffer (50 mM phosphate, pH 7.8), containing EDTA (1 mM) and riboflavin-NBT in the dark for 10 min. The riboflavin-NBT was replaced by 0.1%v/v TEMED and left in the dark for 15 min.
How do you calculate enzyme activity in SOD?
In presence of SOD in reaction mixture, NBT will produce less amount of coloured complex than control. % inhibition of NBT reduction by SOD = control OD- treatment OD/ control x 100 =X% inhibition. 50% inhibition is equal to 1 unit of enzyme. then X% is equal to 1/50 x X= Y unit.
What is the function of catalase and superoxide dismutase?
Superoxide dismutase (SOD) and catalase are enzymes that protect cells from radical attack. Catalase disproportionates hydrogen peroxide, and SOD is an oxidoreductase that serves to dismutate the superoxide anion.
What is SOD assay?
The Cayman Chemical SOD Assay kit is a fast and reliable assay for the measurement of SOD activity from plasma, serum, tissue homogenates, and cell lysates. SOD activity is assessed by measuring the dismutation of superoxide radicals generated by xanthine oxidase and hypoxanthine in a convenient 96 well format.
What is superoxide dismutase assay?
Our OxiSelect™ Superoxide Dismutase Activity Assay uses a xanthine/xanthine oxidase (XOD) system to generate superoxide anions and a chromagen to produce a water-soluble formazan dye upon reduction by superoxide anions. The superoxide dismutase activity is determined as the inhibition or reduction of chromagen.
What is the unit of superoxide dismutase?
The SOD activity was expressed as U/ml reagent. One unit is the amount of SOD that inhibits the rate of formazan dye formation by 50%.
How do you calculate the dilution factor of an enzyme assay?
If you dilute your enzyme by adding, say, 1 µL of enzyme solution to 4 µl buffer, then you have a 1 / (1+4) = 1/5 dilution. The above experiment gives you the concentration of enzyme in the diluted sample, to get the concentration of the stock you have to divide by the dilution, that is, multiply by 5 (the reciprocal).
What is SOD antioxidant?
Superoxide dismutase (SOD) is the first detoxification enzyme and most powerful antioxidant in the cell. It is an important endogenous antioxidant enzyme that acts as a component of first line defense system against reactive oxygen species (ROS).
How do you calculate SOD?
To figure out how much sod you need, measure out the area; get the length in feet and the depth in feet. Multiply these 2 numbers together to get total square feet. Divide this number by 9 to get square yards. Sod is sold by piece and by square yard (which is 2.25 pieces).
What is the full form of SOD?
|SOD||Safe on Deck|
|SOD||Sediment Oxygen Demand|
|SOD||Stormtroopers of Death (band)|
|SOD||Separation of Duties|
How do SOD and catalase work together to neutralize toxic oxygen?
Presence of enzymes catalase & superoxide dismutase (SOD) allows toxic forms of oxygen to be neutralized; can use oxygen. Presence of one enzyme, SOD, allows harmful forms of oxygen to be partially neutralized; tolerates oxygen.
How do you prepare sod assay tubes?
Using the diluted sample, prepare total SOD assay tubes to contain 2, 5, 10, 15, 25, 50 and 100 μg of protein. Next, using the original lysate, prepare total SOD assay tubes containing 200, 300 and 500 μg protein. Add the necessary amount of PB to each tube for a final volume of 100 μl.
What is the difference between the epinephrine and pyrogallol methods for SOD?
Both methods are capable of detecting and quantifying SOD; under these conditions, the epinephrine method is more sensitive to lower concentrations of SOD, but the pyrogallol method yields more linear results in the high range of SOD concentration.
How do I perform the staining step for SOD activity gel?
All staining steps are performed at room temperature. Prepare and run all other aspects of the assay as for the SOD activity gel (see Box 1, Steps 1 -7). For samples prepared for cell culture, load 200-300 μg protein per sample. For tissue homogenates, load 100-200 μg protein per sample.
Is pyrogallol a suitable method for tissue extraction rich in glutathione?
The pyrogallol method was not significantly affected by large excesses of glutathione and appears to be a superior method for tissue extracts likely to be rich in glutathione. Research Support, Non-U.S. Gov’t