Single Molecule Imaging of Neurotrophin Signaling in Axons
Single molecule measurements show great potential for characterizing complex dynamic behavior: such measurements allow one to look beyond the ensemble average and measure real-time trajectories of individual molecules to determine the exact distributions of molecular properties. In the light of new technique advances, single molecule fluorescence imaging studies have been carried out in live cells and provide a direct way to quantify biological events inside cells with a high spatial and temporal resolution.
We are interested in developing and applying single molecule fluorescence imaging method to visualize signal flow in neurons. The biological question of interest is the molecular mechanisms associated with neurotrophin signaling and the implications of those mechanisms for neurodegenerative diseases. Methods of interest include pseudo total-internal-reflection-fluorescence (TIRF) microscopy, fluorescence-resonance-energy transfer (FRET) nanometer localization of a single fluorophore, and using quantum dots as a novel fluorescent label. Standard biochemical and molecular biology methods (cloning of genes, expression of recombinant proteins in bacteria, and transfection of plasmids into mammalian cells etc.), are used to exam cellular events associated with neurotrophin signaling.
Microfluidic culture platform for neuronal networkThe human brain is a complex network composed of ~ 100,000,000,000 neurons, each making up to ~ 10,000 connections to other neurons. The structure and connection of neural network is at the root of the enormous sophistication and computational power of the brain. Thus, it is of great interest to understand the relation between the neuronal network spatial-organization and its functional activity. We are interested in developing microfluidic platforms capable of cultivating neuronal networks with defined connection pattern and separately controlled chemical environment for pre- and post-synaptic neurons. Multi-layer soft-lithography and surface patterning methods are used to build microfluidic devices. Patterned microelectrodes are used to stimulate and monitor network activities with single cell precision.