Predictive-focus illumination for reducing photodamage in live-cell microscopy |
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Authors: | Schilling Z Frank E Magidson V Wason J Lončarek J Boyer K Wen J Khodjakov A |
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Affiliation: | Department of Electrical, Computer, and Systems Engineering, Rensselaer Polytechnic Institute, Troy, New York, USA. |
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Abstract: | Due to photobleaching and phototoxicity induced by high-intensity excitation light, the number of fluorescence images that can be obtained in live cells is always limited. This limitation becomes particularly prominent in multidimensional recordings when multiple Z-planes are captured at every time point. Here we present a simple technique, termed predictive-focus illumination (PFI), which helps to minimize cells' exposure to light by decreasing the number of Z-planes that need to be captured in live-cell 3D time-lapse recordings. PFI utilizes computer tracking to predict positions of objects of interest (OOIs) and restricts image acquisition to small dynamic Z-regions centred on each OOI. Importantly, PFI does not require hardware modifications and it can be easily implemented on standard wide-field and spinning-disc confocal microscopes. |
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Keywords: | Digital imaging light microscopy live‐cell microscopy photodamage software |
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