The environmental risk assessment of nanoparticles (NPs) relies on the adaptation of toxicity protocols originally conceived for contaminants having their predominant bioavailable form in the soluble one. While protocols to describe NP characteristics, behaviour and transformations in exposure media are available, no substantial update of traditional test guidelines has been proposed concerning toxicity endpoints and tests duration. Besides the eventual release of toxic metal ions, nano-ecotoxicity occurrence necessarily involve the physical contact between NPs and aquatic organisms. This relies on the onset of nano-bio interfacial interactions, driven by the properties of both NPs and biological surfaces. Superficial adhesion to the organism exterior (cell membrane/wall or body surface) and ingestion are usually the first outcomes of NP exposure but are rarely followed by dramatic outcomes in the short-term exposures. However, when prolonged exposures are performed, these side-effects can turn into more severe outcomes, with consequences also at the behavioural and ecological level. This chapter wants to highlight the importance of observing and reporting sub-lethal effects in NP acute exposures, such as adhesion and ingestion, and use them as early-warning signs of toxicity to look for to determine the need to perform a chronic exposure. The complex interplay of factors behind the occurrence of nano-ecotoxicity makes the environmental risk assessment of NP less straightforward compared to that of soluble compounds and may require a dedicated approach. This will help to obtain more environmentally relevant data and avoid the risk of underestimating the potential threat posed by NPs to aquatic ecosystems.