Structured Illumination and Dark-Field Microscopy

Project title: Integrated structured illumination and dark field microscope for Molecular, Metabolic and Vascular Imaging

Project Description: The goal of this project is to integrate dark field microscopy (DFM) and structured illumination microscopy (SIM) into one high throughput imaging system to quantitatively image molecular, metabolic and vascular features in biological systems. The spectral images coupled with optical modeling techniques can be used to quantify oxygen saturation and hemoglobin concentration from absorption imaging and glucose uptake and mitochondrial membrane potential from 2-NBDG and TMRE fluorescence imaging, respectively. Our group has previously developed a structured illumination microscope (SIM), which has been successfully adopted for fluorescence imaging at the cellular level (~4.4 µm) with a single frame FOV of 2 x 1.6 mm2. In an independent effort, our group has also developed a dark field microscope (DFM) with a resolution of 7 µm and a FOV of 1.5 x 1.5 mm2 to image the absorption of tumor vasculature. In this project, the two independent microscopy systems were integrated into one system with a common FOV of 2 x 1.6 mm2 and resolution of ~4.4 µm to quantitatively image the different hallmarks of cancer in a seamless manner. The proposed technology can be potentially used to study tumor bioenergetics and molecular contrast in the context of early detection, invasion, metastatic potential, and resistance to therapy.

(A)Schematic of DFM. The key components in DFM are the stop and iris with optimized size to remove the specular reflectance while maintaining high contrast; (B) Schematic of SIM. The key component in SIM is the spatial light modulate (SLM) that can generate illumination beam with fringe patterns to enhance the resolution and contrast by rejecting out of focus background fluorescence; (C) Schematic of SIM-DFM; The key component in the SIM-DFM is the LCTF in their shared collection channel which make the system have spectral capability for both of absorption and fluorescence imaging. (D) Blood vessel phantom imaging by SIM-DFM; (E) Imaging of 10 µm fluorescent spheres by SIM-DFM.