I am trying to find the enzyme activity of catalase. I have assayed my test enzyme sample alongside my standards (bovine liver catalase). I measured the absorbance at 520 nm and plotted a graph of the standard curve (absorbance vs. units/ml of catalase standard). How can I use this graph and the equation of the line to find the enzyme activity in units/ml for my test enzyme samples?
The standard curve should have a linear portion at the lower end of catalase concentrations, meaning that the absorbance increases in direct proportion to the catalase concentration. The slope of this part of the standard curve has units of absorbance/(U/ml).
Measure the absorbance produced by a range of dilutions of your enzyme sample under identical conditions as for the standard curve, and make a plot of absorbance versus enzyme dilution. By dilution, I mean the fractional concentration of the undiluted sample. For example, if you make a 1:20 dilution, the concentration of the dilution relative to the original sample is 1/20 = 0.05. This curve should also have a linear portion. Measure the slope of the linear portion. The slope has units of absorbance/dilution.
Finally, divide the second slope by the first: (absorbance/dilution)/(absorbance/(U/ml). This will give the U/ml of the undiluted sample.
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