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Nanoparticles can be synthesized in a wide array of shapes and sizes to suit specific biomedical applications in therapy and imaging. Prerequisite to such applications are particle stability in biological environments, non-toxicity, and facile conjugation of the particle surface with targeting biological moieties (such as antibodies). Here we report significant flaws in the common methods used to functionalize the surface of gold nanorods (GNRs) of larger-than-usual sizes. We find that while GNRs of sizes smaller than 50 x 15 nm can be effectively stabilized by polyethylene glycol (PEG)-based methods, larger GNRs form major aggregates and crash under similar functionalization conditions. Large GNRs may provide enhanced imaging sensitivity in biological applications due to greater optical extinction cross sections, provided that the GNRs can be made biostable. In this study, GNRs of sizes up to 90 x 30 nm were synthesized using two different published methods. Particle morphology and size distributions were characterized using Transmission Electron Microscopy (TEM), and optical spectra were measured by Vis-NIR Spectrometry. The colloidal stability of different-sized GNRs was assayed at various stages of functionalization using zeta potential and Vis-NIR measurements. The results of these experiments indicate that large GNRs functionalized with PEG undergo irreversible aggregation after minimal washing. We find that coating large GNRs with polystyrene sulfonate (PSS) instead of PEG vastly improves GNR stability in water and serum. Moreover, we provide a novel platform for conjugating biomolecules of interest to PSS-coated large GNRs. We show that larger GNRs produce stronger photoacoustic signal than commonly used smaller GNRs, indicating an advantage of using large GNRs for biomedical imaging. Our observations underscore that the biomedical advantages of novel nanoparticle synthesis methods may not be realized without tailored surface functionalization methods. More generally, our results suggest that materially-identical nanoparticles (i.e. GNRs) exhibit varying stability as a function of particle size.
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