Another interesting aspect of coupled enzyme catalyzed reactions with a zymogen activation step (1)-( ) is the quantification of the catalytic conversion of zymogen in vitro. Formally, the quantification of enzyme activity through measurements obtained from an in vitro assay is mathematically known as an inverse problem. If the activation step (1) is not detectable experimentally (i.e., non-observable), then the secondary reaction step (2) is selected to be an easily observable reaction. This is done with the goal of measuring the enzyme activity of the non-observable reaction through analysis of progress curves generated by the observable reaction. In this case, the secondary reaction step (2) is known as the indicator reaction. Traditionally, coupled enzyme assays are designed so that the product of the non-observable reaction is a substrate for the secondary enzyme in the indicator reaction (see for specific applications). While this type of assay is well-studied , in vitro assays that consist of a zymogen activation step have not been analyzed with the same degree of interest.