Manufactured nanoparticles (NPs) belong to an emerging class of potential environmental pollutants. Of particular interest are the characteristics of NP toxicity under different exposure conditions e.g. cell culture, aquatic or soil media. NPs are thought to behave differently depending on the media making it challenging to establish links between cause and effect. For example, natural organic matter such as humic acid is known to interact with inorganic NPs changing the surface properties and thus their colloidal stability, which cannot be seen in deionised water media. Earthworms are versatile indicator organisms that can be tested in a wide range of different exposure media providing the possibility for a mechanistic insight into exposure testing of NPs. Here we report characterisation of Ag-NPs and their toxicity to the earthworm Eisenia fetida based on soil and aquatic exposures as well as direct injection. The details of challenges regarding the methodology, including stabilisation of Ag-NPs in physiological salt solutions, are discussed. Common problems in discriminating release of free ion Ag+ from Ag-NP in environmental media and in vivo are also addressed.