Quantifying changes in the optical spectra of plasmonic nanoparticles following interaction with biological cells: Towards optimized design in biomedical applications
Chen, Allen L
Drezek, Rebekah A
Doctor of Philosophy
As plasmonic nanoparticle (NP)-based diagnostic and therapeutic technologies—such as Plasmon Resonance Energy Transfer (PRET)-based intracellular analysis or NIR photothermal cancer therapy—continue to be developed with the goal of eventual clinical translation, clearer understanding of the interactions between NPs and biological environments is critical. In biological media, proteins adsorb to NPs, leading to physicochemical changes which affect the cellular uptake of NPs, and NPs can further agglomerate within intracellular vesicles. Since the plasmonic properties of NPs are highly dependent on their geometry and local environment, these physicochemical changes can also change the optical spectra of NPs in cellular environments. Understanding how NPs’ optical properties change in biological environments is especially important as medical nanotechnologies move toward increased multiplexing capabilities, and shifted optical spectra may result in unintended outcomes. In this thesis, we develop a darkfield hyperspectral (HS) imaging approach for systematically studying and quantifying how the NP optical spectra change in a cellular environment in order to inform biomedical NP design. We begin by establishing methods for measuring and analyzing spectra of plasmonic NPs in a cellular environment, showing that 100 nm gold NPs (AuNPs) experience up to a 79 nm spectral shift and substantial spectral broadening after exposure to Sk-Br-3 breast adenocarcinoma cells for 24 h. Then, we apply this HS imaging approach to characterize how NP design factors and biological environment factors impact the extent of spectral changes experienced by NPs within cellular environments. We find that NP functionalization with poly(ethylene glycol) (PEG) can reduce spectral shifting and decrease spectral broadening exhibited by NPs upon cellular uptake, and we show the impact of serum concentration on the magnitude of spectral shift exhibited by NPs. Finally, in order to more specifically focus characterization on the optical response from cell-internalized NPs and improve understanding of intracellular delivery of NPs, methods are needed to differentiate membrane-adsorbed NPs from cell-internalized NPs. Using confocal imaging, we investigate the efficacy of emerging approaches for differentiating cellular internalization from cell membrane adsorption of NPs to enable further advances in understanding of nano-bio interactions. Together, this thesis demonstrates the development of a HS imaging and analysis approach for quantitatively characterizing changes in the optical properties of NPs within cellular contexts, and the application of this approach towards developing quantitative relationships to enable plasmonic NPs to be engineered to exhibit desired optical properties in biomedical environments for maximal efficacy and safety.