Quantitative Label-Free Imaging of Nanoparticle-Cell Interactions Using Light-Scattering-Based Widefield Microscopy

Abstract Nanoparticle-cell interactions often determine the safety and efficacy of nanomedicines. To visualize these interactions with optical microscopy, researchers typically label nanoparticles fluorescently. However, such labeling can alter nanoparticle-cell interactions by changing nanoparticle surface chemistry. Here, we describe a label-free, light-scattering-based imaging workflow that uses a conventional widefield fluorescence microscope to study nanoparticle-cell interactions. To demonstrate proof of concept, we custom-designed several light-filter cubes that enable sensitive detection of nanoparticle scattering signals. We used gold nanoparticles as a model system and demonstrated up to a ∼78-fold increase in the median light-scattering signal by varying nanoparticle surface chemistry. We further showed the suitability of this imaging approach for live-cell tracking, with no measurable phototoxicity after 6 h of continuous illumination, enabling long-term studies critical for nanomedicine development. Our accessible, cost-effective workflow enables researchers to conduct longitudinal, label-free, and quantitative studies of nanoparticle-cell interactions without the need for specialized microscope systems.