Soutenance de these Corey BUTLER "Quantitative Single Molecule Imaging Deep In Biological Samples Using Adaptive Optics"

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Soutenance de thèse Corey BUTLER

 

"Quantitative Single Molecule Imaging Deep In Biological Samples Using Adaptive Optics"
 
Résumé:
Optical microscopy is an indispensable tool for research in neurobiology and medicine, enabling studies of cells in their native environment. However, subcellular processes remain hidden behind the resolution limits of diffraction-limited optics which makes structures smaller than ~300nm impossible to resolve. Recently, single molecule localization (SML) and tracking has revolutionized the field, giving nanometer-scale insight into protein organization and dynamics by fitting individual fluorescent molecules to the known point spread function of the optical imaging system. This fitting process depends critically on the amount of collected light and renders SML techniques extremely sensitive to imperfections in the imaging path, called aberrations, that have limited SML to cell cultures on glass coverslips.

A commercially available adaptive optics system is implemented to compensate for aberrations inherent to the microscope, and a workflow is defined for depth-dependent aberration correction that enables 3D SML in complex biological environments. A new SML technique is presented that employs a dual-objective approach to detect the emission spectrum of single molecules, enabling 5-dimensional single particle imaging and tracking (x,y,z,t,λ) without compromising spatiotemporal resolution or field of view. These acquisitions generate ~GBs of data, containing a wealth of information about the localization and environment of individual proteins. To facilitate quantitative acquisition and data analysis, the development of biochemical, software and hardware tools are presented. Together, these approaches aim to enable quantitative SML in complex biological samples.

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  • Nom : BUTLER Corey