In 3ml, how can I make all these buffers, I have read this paper for SOD activity, Please find below
[The 3 ml reaction mixture contained 50 mM potassium phosphate buffer (pH 7.0), 0.5 mM ascorbic acid, 0.1 mM EDTA, 1.5 mM H2O2 and 0.1 ml enzyme]
Any one can help me?
It is easy. It depends in how many samples you have. For example, you have 30 samples, which means that you need 90 ml buffer totally. You can prepare 150 ml buffer. According to the concentration of each component, you can calculate the amount of each product. And finally, all the products are dissolved in 150 ml water. Before up to 150ml, don't forget to adjust the pH in advance.
Anyway, it is impossible to prepare 3 ml each time. It is funny......
Briefly, 100 µl enzyme, 100 µl of 186 µM phenazene methosulfate, 300 µl of 3.0 mM nitroblue tetrazolium, 200 µl of 780 µM NADH were incubated for 90 seconds at 30°C. The reaction was terminated by adding 1000 µl of acetic acid and the addition of 4 ml n-butanol. The absorbance was recorded at 560 nm using a UV-VIS Biospectrophotometer. The percent inhibition was calculated by measuring the absorbance of blank without SOD enzyme and the test with SOD enzyme samples using the formula (Blank-Sample)/Blank X 100.
you can try this protocol if you want
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