Standard analyses of enzyme kinetics (the only kind discussed here) assume:

•The production of product is linear with time during the time interval used.

•The concentration of substrate vastly exceeds the concentration of enzyme. This means that the free concentration of substrate is very close to the concentration you added, and that substrate concentration is essentially constant throughout the assay.

•A single enzyme forms the product.

•There is negligible spontaneous creation of product without enzyme.

With the exception of the built-in allosteric model, Prism's built-in equations also assume:

•No cooperativity. Binding of substrate to one enzyme binding site doesn't influence the affinity or activity of an adjacent site.

•Neither substrate nor product acts as an allosteric modulator to alter the enzyme velocity.