Remane’s Artenminimum (species minimum) at the horohalinicum (5 - 8 psu, i.e., in brackish water) is a fundamental concept in ecology to describe and explain the distribution of organisms along salinity gradients. In stark contrast, for protists, a recent metadata analysis challenged this concept and proposed a species maximum in brackish waters. This literature-based investigation was however criticized because of the overrepresentation of brackish water studies. Reliable data to verify or reject the species minimum for protists in the critical salinity zone connecting freshwater and marine habitats are thus critically lacking. In this study, pronounced horizontal (west-east transect) and vertical (Kiel Bight, Arkona Basin, Bornholm Basin) salinity gradients in the Baltic Sea with an even coverage of different salinity classes were sampled and protistan plankton communities were analyzed using high-throughput eDNA metabarcoding. The question was addressed on how salinity gradients shape autochthonous brackish protistan plankton communities. The objectives were to obtain samples allowing unbiased analyses of alpha-diversity patterns along a salinity gradient, to then better relate protistan community shifts to changes in salinity and other potential environmental drivers both along horizontal and vertical gradients, and ultimately to test the validity of Remane’s Artenminimum concept for planktonic protists.