A detached leaf disk assay for screening sweet cherry (Prunus avium L.) genotypes for susceptibility to powdery mildew (PM) [Podosphaera clandestina (Wallr.:Fr.) Lev.] was developed by evaluating the effects of photoperiod (24 hours light, 0 hours light, 14 hours light/10 hours dark), substrate nutrient content (sterile distilled water, 1% sucrose), leaf age (old, young, emergent), and leaf explant size (intact leaf, 30 mm, 20 mm) on PM growth on leaves from the susceptible cultivar Bing. The only parameter described that had a significant (P ≤ 0.001) effect on PM growth was leaf age. Old leaves, designated as the third fully expanded leaf from the basal end of current-year's shoot growth, were never infected with PM under controlled inoculations. In the absence of significant differences between treatments, those parameters with the highest treatment means were selected for subsequent evaluation. To test the leaf disk assay, 14 sweet cherry cultivars were screened in two experiments, and rated according to level of PM susceptibility. Rank sum comparison of results from cultivars used for leaf disk screening agreed with earlier field rankings of the same cultivars. The developed leaf disk assay greatly reduced the space required to screen sweet cherry cultivars, and was a repeatable and objective predictor of field resistance that may be useful for screening germplasm or breeding populations.