High Tempo-spatial Resolution Single Particle Microscopic Imaging

The unique localized surface plasmon resonance from noble metal nanoparticles facilitates the transduction of chemical or physical stimulus into optical signals in a highly efficient way. With the development of high-throughput and nondestructive optical microscopic technology,we can discuss the important chemical and biological processe at single cell and single molecule level.Further development and application of optical imaging methods based on plasmonic probes should lead to many exciting results in chemistry and biology in the future.Based on the challenges put forward in previous reports,the work in this thesis was performed as follow:1. With time-resolved high-precision single-particle tracking methodologies,we explored the adsorption and thermal motion of transacting activator of transcription (TAT) peptide-modified nanocargo on a model lipid bilayer in the nonelectrostatic domain.We found that the lateral and rotational motion of TAT peptide-modified nanocargo could be effectively suppressed on the surface of neutral lipid membrane,a feature that cannot be explained by existing hypotheses.A semiquantitative association activation energy analysis revealed that multiple weak bonds were required for the initial adsorption process.As a result, the localized multiple TAT peptides on the surface of the nanocargo can provide a pathway for the creation of a net of peptide—ipid complexes (e.g., lipid domain).The dragging forces caused by these complexes effectively confined the thermal motion of the nanocargo on thefluid membrane that cannot be achieved by individual peptides with random spatial and conformational distributions.These interesting findings could provide insightful information for the better understanding of the intracellular internalization mechanism of TAT peptide-modified nanocargo and might shed new light on the development of highly efficient intracellular carriers for site-specific delivery of drugs and genes.2. In this work, with a dual wavelength view darkfield (DWVD) microscope, the tempo-spatially resolved dynamics of TAT peptide-functionalized gold nanoparticles (TGNPs) in living HeLa cells were explored. It was found that energy-dependent endocytosis was the prevailing pathway for the cellular uptake of TGNPs.Active nuclear targeting was not observed according to the time-correlated dynamic spatial distribution information.Interestingly, inheritage of TGNPs to the daughter cells through mitosis was found to be the major route to metabolize TGNPs by HeLa cells.These understandings on the cellular uptake mechanism and intracellular fate of nanocargos in living cells would provide deep insight on how to improve and controllably manipulate the translocation efficiency for targeted drug delivery. 3. Many intriguing physical and chemical processes are taken place at liquid-solid interface.Total internal reflection illumination together with single molecule spectroscopy provides a robust platform to selectively explore kinetic processes close the interface.With this technique, we demonstrated that the quantity of Rhodamine B molecules close to a chromatography surface could be regulated in a photo-driven way. It was found that laser-induced repulsion force at this interface could form an energy barrier in a long-range fashion.The strength is enough to compromise the thermal diffusion of single molecule in a range of several hundred nanometers normal to the liquid-solid interface.Therefore,it might display extensive applications in the development of photon modulation technique with high throughput capability and applies filters of different sizes nanoparticles.