## 3 Characterisation
### Sample Preparation  
Many NMs are supplied as powders and need proper dispersion in vehicle media to allow the assessment of their human and environmental hazards [43]. The characterization of NMs in stock dispersions in ultrapure water or a physiological buffer is well-established and researchers generally agree on how to do this and what to measure. But, this agreement is harder to achieve for complex media (natural water, soil, cell culture media with serum proteins, and blood samples) because the NMs behave differently in each milieu, such as agglomeration or formation of a protein corona changing their surface reactivity and size. The addition of serum, albumin of detergents to allow a better and more stable dispersion could also modify the biological responses induced [44]. One solution to such sample preparation problems is to have a decision-tree about characterization based on what is possible in each type of media, and relevant for the exposure route and hazard end-point being assessed. A good dispersion without modification of NM characteristics, especially surface reactivity due to coating with proteins or other biomolecules, is necessary to mimic accurately real life exposures. This preparation of NMs should take into account the exposure routes and target organs to be studied. Thus during inhalation the nanoparticles will come into contact with lung lining fluid mainly composed of lipids, whereas ingested nanoparticles will encounter the acidic gastrointestinal fluids. Standardized dispersion protocols are established or under development to harmonize preparation methods (OECD guidelines) but their impacts on biological responses still need further investigation. Despite considerable progress, there is continuing uncertainty around specific issues such as the role of the synthesis route on the surface composition of nominally identical NMs. For example, silica nanoparticles can be prepared by at least five different routes, utilising different catalysts, _etc._, with the consequent emergence of subtle differences in the surface composition. Currently, researchers (and industry) do not distinguish between these as potentially different NMs, with potentially different uptake and impacts, and as such the published data are not always reliable and comparable. [45; 46]  
A key communication issue is that authors are not reporting characterisation _during_ the exposures, either because of logistical hurdles or because the technology is simply not reliable in the media and not with a low enough detection limit. **Recommendation 12**: We recommend that a pragmatic way forward is for manufacturers to cooperate in characterizing the relevant properties of the NMs at production (see [PERSON] _et al._, 2010 for further details [10]), and to then further explore a very limited set of parameters before use (or exposure), e.g., size and size distribution, zeta potential and adsorbed species in order to take into account the effects of storage and sterilization and how different environments/conditions affect the physicochemical characteristics of NMs [4]. **Recommendation 13**: Industry, regulators and standards organizations should cooperate in generating a nomenclature system for NMs that accounts for synthesis route and potentially for surface description including initiator/catalyst residues and potential surface impurities or defects. If this proves to be too challenging, perhaps because there are too many variables, the solution may be to stick with the current chemical nomenclature and additionally indicate particle size and shape [10], adding to the chemical product labelling a code number for the synthesis method. Like other chemicals they also need to list major impurities that relate to the synthesis. **Recommendation 14**: It is good practice for any assay to be cross-checked for interference, and the authors also emphasize that NM samples could interfere with the test assays themselves. We recommend the establishment of appropriate controls for such possible interference [7].