Open AccessMiniature scanning light-sheet illumination implemented in a conventional microscope
Author(s) -
Anjan Bhat Kashekodi,
Tobias Meinert,
Rebecca Michiels,
Alexander Rohrbach
Publication year - 2018
Publication title -
biomedical optics express
Language(s) - Uncategorized
Resource type - Journals
SCImago Journal Rank - 1.362
H-Index - 86
ISSN - 2156-7085
DOI - 10.1364/boe.9.004263
Subject(s) - light sheet fluorescence microscopy , optics , materials science , microscope , optical sectioning , lens (geology) , confocal , optical axis , microscopy , optical tweezers , bessel beam , beam (structure) , scanning confocal electron microscopy , physics
Living cells are highly dynamic systems responding to a large variety of biochemical and mechanical stimuli over minutes, which are well controlled by e.g. optical tweezers. However, live cell investigation through fluorescence microscopy is usually limited not only by the spatial and temporal imaging resolution but also by fluorophore bleaching. Therefore, we designed a miniature light-sheet illumination system that is implemented in a conventional inverted microscope equipped with optical tweezers and interferometric tracking to capture 3D images of living macrophages at reduced bleaching. The horizontal light-sheet is generated with a 0.12 mm small cantilevered mirror placed at 45° to the detection axis. The objective launched illumination beam is reflected by the micro-mirror and illuminates the sample perpendicular to the detection axis. Lateral and axial scanning of both Gaussian and Bessel beams, together with an electrically tunable lens for fast focusing, enables rapid 3D image capture without moving the sample or the objective lens. Using scanned Bessel beams and line-confocal detection, an average axial resolution of 0.8 µm together with a 10-15 fold improved image contrast is achieved.