The application of label-free Raman microscopy to monitor particle-cell interactions in in vitro experiments

Characterizing particle-cell interactions is a critical component of nanotoxicology research, complementing material characterization to better interpret observed biological effects and explore dose-response relationships. However, accurate assessment of particle uptake remains challenging due to limitations in distinguishing internalized particles from those bound to the cell surface, labor-intensive sample preparation, and constraints in quantification methods. To address these challenges, label-free techniques with minimal sample processing are being explored. In this study, we present findings from a series of experiments using confocal Raman microscopy as a non-destructive, label-free method to evaluate particle-cell interactions. Suspensions of anatase (NM-102; 21 ± 10 nm) and rutile (NM-104; 26 ± 10 nm) TiO₂ nanoparticles were analyzed in two in vitro models: THP-1 (suspension) and 16HBE14o- (adherent) cells. Raman spectral analysis enabled the detection of cell-associated particles based on their unique chemical fingerprints. We further observed concentration-dependent trends in particle-cell interaction using semi-quantitative metrics such as particle area and spectral match. Lastly, optimal scan parameters were identified to improve detection efficiency while preserving simple sample preparation workflows.