The acceleration of nanomaterials research has brought about increased demands for rapid analysis of their bioactivity, in a multi-parametric fashion, to minimize the gap between potential applications and knowledge of their toxicological properties. The potential of Raman microspectroscopy for the analysis of biological systems with the aid of multivariate analysis techniques has been demonstrated. In this study, an overview of recent efforts towards establishing a 'label-free high content nanotoxicological assessment technique' using Raman microspectroscopy is presented. The current state of the art for cellular toxicity assessment and the potential of Raman microspectroscopy are discussed, and the spectral markers of the cellular toxic responses upon exposure to nanoparticles, changes on the identified spectral markers upon exposure to different nanoparticles, cell death mechanisms, and the effects of nanoparticles on different cell lines are summarized. Moreover, 3D toxicity plots of spectral markers, as a function of time and dose, are introduced as new methodology for toxicological analysis based on the intrinsic properties of the biomolecular changes, such as cytoplasmic RNA aberrations, protein and lipid damage associated with the toxic response. The 3D evolution of the spectral markers are correlated with the results obtained by commonly used cytotoxicity assays, and significant similarities are observed between band intensity and percentage viability obtained by the Alamar Blue assay, as an example. Therefore, the developed 3D plots can be used to identify toxicological properties of a nanomaterial and can potentially be used to predict toxicity, which can provide rapid advances in nanomedicine. Graphical Abstract Spectral markers of cytotoxicity as a function of time and dose.
Keywords: Cytotoxicity; Genotoxicity; High content analysis; Nanotoxicology; Raman microspectroscopy; Spectral markers.