We will be conducting an enzymatic assay using a-glucosidase and a-amylase using acarbose as the positive control. What should be our negative control?
One way to do it would be to simply leave out the enzyme, replacing it with the same volume of the buffer you would have diluted it with.
This is not always practical, however, especially when using automated systems to dispense the liquid into assay plates. In that case, you need a potent inhibitor that can be added at a high enough concentration to produce complete inhibition. The identity of potent inhibitors of your particular enzymes can be sought in the literature, or in a database such as BRENDA (https://brenda-enzymes.org/). The problem may be obtaining these substances if they are not catalog items.
Add stopping reagent to assay buffer (generally shifting pH to inactive the enzymes) which includes acarbose also as substrate. Next, add the enzyme to mixture to generate a negative control...This represents matrix related impurities excluding enzyme activity and better activity differentiation can be reported...
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