The aim of this study was to determine the minimal number of particles required for assessing a reliable particle size distribution from images acquired using microscopy. The proposed methodology used the one-sample Kolmogorov-Smirnov statistic to choose an a priori number of particles and the two-sample Kolmogorov-Smirnov test to validate the sub-sampling procedure inherent to microscopic preparation. The methodology was applied to number-based particle size distributions of starch granules.