In this application, we propose to demonstrate the feasibility of spectral domain phase microscopy (SDPM) as a new tool for non-invasive, non-contact, far-field optical characterization of static and dynamic nanoscale structures including living cell membranes. SDPM is a functional extension of spectral domain optical coherence tomography which allows for the detection of structural surface profiles, motions and dynamics with nanometer-scale sensitivity in the axial dimension in real time. Nanometer-scale sensitivity in the axial direction is obtained by using a newly developed implementation of exquisitely phase stable self-referencing interferometry, while maintaining the non-contact advantages of working in the optical far field. Lateral resolution in this technique is limited by physical optics to the micrometer scale. Although our proposed technique has many potential applications in static and dynamic structural analysis of living and non-living nanoscale materials and machines, in this R21 application we concentrate primarily on initial SDPM technology development and feasibility demonstrations of its applications in cell biology. The particular cell biology questions we propose to address with this technology include measuring mechanical activity in developing cardiomyocytes and assessing the magnitude and kinetics of cell migration in neural crest cells.
The specific aims of this proposal are as follows: 1. Develop novel technology implementations for high-speed temporally- and spatially- resolved nanoscale axial displacement measurements using spectral domain phase microscopy (SDPM); 2. Develop signal and image processing approaches for improving the accuracy of SDPM displacement quantitation; 3. Apply temporally and spatially-resolved SDPM for static and dynamic nanoscale measurements in living cells. ? ? ?
