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Microbial digestive enzymes in soil and litter have been studied for over a half century, yet the understanding of microbial enzymes as drivers of ecosystem processes remains hindered by methodological differences among researchers and laboratories. Modern techniques enable the comparison of enzyme activities from different sites and experiments, but most researchers do not optimize enzyme assay methods for their study sites, and thus may not properly assay potential enzyme activity. In this review, we characterize important procedural details of enzyme assays, and define the steps necessary to properly assay potential enzyme activities in environmental samples. We make the following recommendations to investigators measuring soil enzyme activities: 1) run enzyme assays at the environmental pH and temperature; 2) run proper standards, and if using fluorescent substrates with NaOH addition, use a …