With the rapid development of nanotechnology products, there is a significant concern on the adverse effects that might be associated with them. Traditional biological assays are typically used to asses the toxicity in vitro. There are, however, questions regarding the suitability of these assays for this purpose, mainly due to the potential interaction of the particles with the utilized dyes. In addition, this process can be costly and time consuming, as a large number of different assays have to be used. To address some of these issues, Raman spectroscopy is used in this study to investigate the particle-cell interactions. The spectrum of a living cell is a very complex and rich collection of data directly related to its chemical composition. To enhance the data resolution and make the detection of toxicity more robust, data-mining techniques have been deployed. Furthermore, data-mining techniques enable full automation of the entire process, minimizing user input. The Raman spectroscopy successfully evaluated the toxicity of TiO2 nanoparticles by both the peak-by-peak analysis and with the implementation of support vector machines. The particles were found to display cytotoxicity after 36 h of exposure. The results were confirmed by MTT (3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide) assay and are in agreement with the existing literature on the subject. Overall, Raman spectroscopy appears to be among the very few techniques that exhibit low levels of interferences (obscuration, fluorescence, emission, etc.) from the particle addition. Since it does not rely on biomarkers, it can be used in situ for an extended period with minimal effects on the cellular biochemistry. Copyright © 2011 John Wiley & Sons, Ltd.