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Acquisition of a Perkin Elmer Opera Phenix Microscope

We intend to procure a Perkin Elmer Opera Phenix high-throughput spinning-disk confocal imager system for the Protein Imaging Platform at CPR. This microscope is specialized for ultra-sensitive, ultra-fast, automated, three-dimensional confocal microscopy of fixed and live cells, which is a microscope technique very high in demand in the Imaging Platform. The microscope will cover needs for both high-throughput screening and multi-well plate-based live cell imaging.</p>

The Novo Nordisk Foundation Center for Protein Research was established in 2007 at the Faculty for Health and Medical Sciences, University of Copenhagen. The research center promotes basic and applied discovery research on human proteins of medical relevance. CPR has been awarded a major grant from the Novo Nordisk Foundation in 2019 after a comprehensive performance evaluation to enter a new stage of ambitious research, which also includes advanced imaging applications. We intend to procure a Perkin Elmer Opera Phenix high-throughput spinning-disk confocal imager system for the Protein Imaging Platform at CPR. This microscope is specialized for ultra-sensitive, ultra-fast, automated, three-dimensional confocal microscopy of fixed and live cells, which is a microscope technique very high in demand in the imaging platform. The microscope will cover needs for both high-throughput screening and multi-well plate-based live cell imaging.

The Opera Phenix microscope uniquely combines all the technical features that we require for ultra-fast, ultra-sensitive 3D imaging of living cells in a high-throughput fashion. It is the combination of very efficient water immersion objectives, to collect maximum light from weakly fluorescent cells, with highest xyz resolution, the simultaneous recording of color channels enabled by the highly sensitive, multi-camera set-up and proprietary Synchrony Optics, and the very fast and robust autofocus that minimizes exposure of live cells to laser light. We need this maximal speed and sensitivity for our cellular imaging because we use state-of-the-art, Crispr-Cas9-based technology for tagging cellular proteins endogenously with GFP/RFP without over expression. The levels and fluorescence intensity of such endogenously tagged proteins in cells is often very low compared to conventional over expression model systems and require the highest sensitivity from microscopes. Specifically, 1 key experimental application at our institute is the detection of GFP/RFP-proteins in tiny, sub-nuclear spots that represent single repair sites of DNA breaks. Detection and optical separation of those spots requires highest xy resolution that is optimally provided by the proprietary 40x and 60x proprietary water immersion objectives. The microscope utilizes a PreciScan (automated pre-scan re-scan) tool in x, y and z dimensions. This smart image acquisition mode to capture and re-find rare type of cells/phenotypes minimizes the volume of 3D image data by up to 40-fold. The Opera Phenix meets all required demands for our specific cellular imaging applications. This microscope employs a micro-lens enhanced dual Nipkow spinning disk with proprietary Synchrony Optics (US Patent 9.612,428). This enables ultra-fast simultaneous multicolor imaging using the two cameras. This unique feature is important for our imaging applications because it minimizes phototoxicity when we need to take many 3D images of living cells. The microscope has automated image alignment using a quasiperiodic grid, which generates perfectly superimposed images during simultaneous imaging with multiple cameras (US Patent 9,852,864 B2). This enables automatic alignment of multicolor images for error-free and time-saving image processing procedures. The same quasiperiodic grid technology is crucial to re-find the same field with extreme accuracy during time lapse recording where the system will return to the same point over and over again. The Opera Phenix is equipped with proprietary high NA automated water objectives enabling shorter measurement times and meet the required x ,y, z resolution in 3D imaging for our experiments which is very important for our long-term live cell time lapse recordings where automatic water-immersion high NA objectives critically shorten exposure times of cells to phototoxic light. Water immersion objectives are optimally matched to the refractive index of aqueous imaging cell culture medium. This technology is covered by two patent families, granted EP1386189 B1, US7304793 B2, and EP1646902 B1, US7961384 B2. The above mentioned PreciScan imaging mode to selectively record rare cells with specific features, uses proprietary image analysis algorithms (STAR) and built-in machine learning for cell classification (called Phenologic). Patents around this STAR method: WO2013068781, WO2013068780, WO2013038225. Patents around the PhenoLOGIC classification method: WO2013038224.

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