Methods and kits are provided for measuring protease activity in samples such as feed or food samples. The methods include adding a water insoluble substrate with a signal producing group to a feed or food sample containing the protease activity to be measured, incubating the sample in phosphate-free buffer such that the signal is produced, and measuring the amount of protease activity in the sample. The methods do not require separation of the incubation buffer from the feed or food and allow for visual inspection of a colorimetric signal for a semi-quantitative measurement of the in-feed/in-food protease activity. Thus, the assay is advantageous as it can be performed on-site without the use of laboratory equipment.
