The objective of this study was to develop an instrument for measuring the quality of enzymatic detergents and analyze their cleaning action on surgical instruments. The instrument was submitted to a panel of three judges. It was pre-tested in a sample of 25 surgical instruments. Twenty-five other surgical instruments trays were used for the field analysis. The results made possible the construction of an instrument with 19 items, and the field analysis verified that there were differences in the scores of the solutions that were evaluated. P5 had the best average score, 78.7 (SD 6.68), and median of 80.2, and P4 had the worst performance, with an average score of 56.2 (SD 7.09) and median of 56.7 in a possible range between 19 and 95. In the study, the measuring instrument proved to be reliable, with a Cronbach's alpha of 0.83.