Ramona Optics Inc

A Parallelized Computational Microscope Platform for High-throughput Live Imaging of Patient-derived Organoids - Significance: as a Biologically Relevant Miniaturized Model System for Human Tissues, 3D Organoids Offer a Powerful New Platform for High-throughput Biomedical Research, Including for the Discovery of Novel Cancer Treatment Regiments. While Microscopy Plays a Vital Role in Organoid Experimentation, Current Imaging Systems Are Inefficient at Capturing Their 3D Morphology in Large Numbers, Which Is Required by a Wide Variety of High-throughput Experiments. in Addition, Current Microscopes Are Unable to Record 3D Video of Dynamics, Such as Cell Viability, Cell Migration, and Processes That Can Reveal Mechanistic Properties Like Diffusion, Permeability, and Organoid Compaction, Across More Than Just a Few Samples in an Effective Manner – Resulting in Experimental Challenges Across a Wide Variety of Contexts. Proposal: Ramona Optics Aims to Develop a New Computational Microscope Platform (MCAM-96) for Parallelized High-content 3D Imaging of Volumetric Specimens, Such as 3D Tumor Organoid Models, at 50X Faster Speeds Than Current Devices Across Entire Well Plates. the MCAM-96 Will Also Record Parallelized Live-cell Videos Assays That Probe Novel Mechanistic and Dynamic Processes. Jointly Developed Open-source Image Analysis Software Will Rapidly Compute Temporally Aligned Statistics of Organoid Morphology as a Function of Time. We Will Benchmark System Performance in a Series of Novel Live Organoid Imaging Experiments at Two Sub-award Labs in Preparation for Our First Market-ready Product. SA1: MCAM-96 Hardware and Software Development and Verification: Ramona Optics Will Produce New Parallelized Microscope Hardware to Simultaneously Image 96 Full Wells (8X12 Array) With 2 Fluorescence Channels at 0.3 ΜM and 4 ΜM Half-pitch Lateral and Axial Resolution, Across a 100 ΜM Axial Range in 10-45 Sec. Parallelized Software Will Register and Calibrate All 96 3D Scans for Database Upload in 15 Sec. We Will Benchmark System Performance Against a Matching Step-and-scan Microscope With 10 Unique 96 Well Plates Containing a Variety of Fluorescently Labeled 3D Tumor Organoids. SA2: Measuring the Cytotoxic Effects of Chemotherapy in Liver Organoids: We Will First Develop Software to Crop Individual Organoids, Project 3D Imagery to 2D In-focus Composites, and Subsequently Extract 100 Key Morphological Features Per Organoid Per Scan. With the Rajagopalan Lab at Virginia Tech, We Will Then Test Our New System Workflow in a Cell Viability Assay That Longitudinally Monitors the Cytotoxic Effects of Chemotherapy (3X Drugs) on 3D Liver Organoids With Tumor Cells. SA3: Tracking Tumor Organoid Toxicity and Cell Migration During Immunotherapy: With the Soker Lab at Wake Forest, We Will Enhance the MCAM-96 With a Parallelized Tilted Light Sheet Array for Optical Sectioned 3D Capture, Before Completing a Series of Experiments That Record 96X Parallelized Time- Lapse Videos of T Cells Co-cultured With Tumor Organoids During Variable-dose Immunotherapy Treatment.