Poster abstracts
Poster number 23 submitted by Thomas Porter
Photoswitchable quantum dot-gold nanoparticle probes for super-resolution microscopy
Thomas Porter (William G. Lowrie Department of Chemical and Biomolecular Engineering), Abhilasha Dehankar (William G. Lowrie Department of Chemical and Biomolecular Engineering), Kil Ho Lee (William G. Lowrie Department of Chemical and Biomolecular Engineering), Jessica Winter (William G. Lowrie Department of Chemical and Biomolecular Engineering, Department of Biomedical Engineering)
Abstract:
Super-resolution microscopy is a powerful tool for sub-diffraction limit imaging of live biological samples. In stochastic optical reconstruction microscopy (STORM), photoswitchable probes stochastically switch between on/off states, enabling individual particles to be imaged exclusive of their nearest neighbors. However, fluorescent dyes currently used limit the achievable resolution of STORM because they have low photon emission rates and are susceptible to photobleaching. In comparison, semiconductor nanoparticles known as quantum dots (QDs) possess greater stability against photobleaching and higher photon emission rates. Furthermore, their size-dependent, narrow emission spectra and broad absorption spectra are ideal for color multiplexing. However, QDs are not inherently photoswitchable. Previously, our lab developed QD-gold nanoparticle (AuNP) probes linked by azobenzene modified complementary ssDNA, where photoswitching was accomplished through Förster resonance energy transfer (FRET). By modulating excitation wavelength, the azobenzene groups photoisomerize between cis and trans conformations, altering DNA duplex stability. By regulating the DNA hybridization state, FRET quenching efficiency is altered through the changing QD and AuNP interparticle distance. Although photoswitching was achieved in this initial model, it did not provide adequate consistency and contrast between on/off states. For FRET-based STORM imaging probes, >95% quenching should be achieved. In this work, we develop a compact QD surface coating using the “loops-trains-tails” polymer adsorption model to minimize interparticle distance. Then, using EDC/sulfo-NHS crosslinker chemistry and click chemistry, we conjugate ssDNA to QDs. As an alternative, we avoid difficulties with QD bioconjugation by directly embedding DNA in the shell of the QD core/shell structure. Here, we report our nanoparticle synthesis techniques and results, as well as the quenching efficiencies of our QD-AuNP probes.
Keywords: Nanotechnology, Nanoparticles, Microscopy