title: Predictive-focus illumination for reducing photodamage in live-cell microscopy.
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.
authors.affiliation: Department of Electrical, Computer, and Systems Engineering, Rensselaer Polytechnic Institute, Troy, New York, USA.
authors.forename: Z
authors.initials: Z
authors.lastname: Schilling
authors.forename: E
authors.initials: E
authors.lastname: Frank
authors.forename: V
authors.initials: V
authors.lastname: Magidson
authors.forename: J
authors.initials: J
authors.lastname: Wason
authors.forename: J
authors.initials: J
authors.lastname: Lončarek
authors.forename: K
authors.initials: K
authors.lastname: Boyer
authors.forename: J
authors.initials: J
authors.lastname: Wen
authors.forename: A
authors.initials: A
authors.lastname: Khodjakov
mesh_terms: D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000379:methods; D008027:Light; D008856:Microscopy, Fluorescence / Q000379:methods*; D038761:Photobleaching / Q000528:radiation effects*; D055213:Retinal Pigment Epithelium / Q000166:cytology* / Q000528:radiation effects