Journal
BIOSENSORS-BASEL
Volume 12, Issue 12, Pages -Publisher
MDPI
DOI: 10.3390/bios12121110
Keywords
light sheet fluorescence microscopy; microfluidic devices; live-cell imaging
Funding
- Ministerio de Ciencia, Innovacion y Universidades, Agencia Estatal de Investigacion [TEC2016-78052, PID2019-109820RB-I00]
- MCIN/AEI
- European Regional Development Fund (ERDF), A way of making Europe
- Ministerio de Ciencia, Innovacion y Universidades, Spain [FPU20/01459]
- Ministerio de Economia y Competitividad [FIS2020-115088RB-I00]
- Horizon 2020 Framework Programme [801347-SENSITIVE]
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This manuscript presents a novel architecture of a single-plane illumination microscopy (SPIM) for high-resolution imaging of live processes at a cellular level. The custom-made microscope overcomes limitations of existing techniques, achieving high-speed acquisition capabilities, low phototoxicity, and low mechanical disturbances, and demonstrating excellent imaging performance in microfluidic devices.
Three-dimensional imaging of live processes at a cellular level is a challenging task. It requires high-speed acquisition capabilities, low phototoxicity, and low mechanical disturbances. Three-dimensional imaging in microfluidic devices poses additional challenges as a deep penetration of the light source is required, along with a stationary setting, so the flows are not perturbed. Different types of fluorescence microscopy techniques have been used to address these limitations; particularly, confocal microscopy and light sheet fluorescence microscopy (LSFM). This manuscript proposes a novel architecture of a type of LSFM, single-plane illumination microscopy (SPIM). This custom-made microscope includes two mirror galvanometers to scan the sample vertically and reduce shadowing artifacts while avoiding unnecessary movement. In addition, two electro-tunable lenses fine-tune the focus position and reduce the scattering caused by the microfluidic devices. The microscope has been fully set up and characterized, achieving a resolution of 1.50 mu m in the x-y plane and 7.93 mu m in the z-direction. The proposed architecture has risen to the challenges posed when imaging microfluidic devices and live processes, as it can successfully acquire 3D volumetric images together with time-lapse recordings, and it is thus a suitable microscopic technique for live tracking miniaturized tissue and disease models.
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