Wrapping nanocrystals with an amphiphilic polymer preloaded with fixed amounts of fluorophore generates FRET-based nanoprobes with a controlled donor/acceptor ratio.
Résumé
Colloidal nanocrystal (NC) donors wrapped with a polymer coating including multiple organic acceptor molecules are promising scaffolds for fluorescence resonance energy transfer (FRET)-based nanobiosensors. Over other self-assembling donor-acceptor configurations, our preloaded polymers have the virtue of producing compact assemblies with a fixed donor/acceptor distance. This property, together with the possibility of stoichiometric polymer loading, allowed us to directly address how the FRET efficiency depended on the donor/acceptor. At the population level, nanoprobes based on commercial as well as custom CdSe/ZnS donors displayed the expected dose-dependent rise in transfer efficiency, saturating from about five ATTO dyes/NC. However, for a given acceptor concentration, both the intensity and lifetime of single-pair FRET data revealed a large dispersion of transfer efficiencies, highlighting an important heterogeneity among nominally identical FRET-based nanoprobes. Rigorous quality check during synthesis and shell assembly as well as postsynthesis sorting and purification are required to make hybrid semiconductor-organic nanoprobes a robust and viable alternative to organic or genetically encoded nanobiosensors.