Dual-beam confocal light-sheet microscopy via flexible acousto-optic deflector

Year: 2019

Authors: Gavryusev V., Sancataldo G., Ricci P., Montalbano A., Fornetto C., Turrini L., Laurino A., Pesce L., de Vito G., Tiso N., Vanzi F., Silvestri L., Pavone FS.

Autors Affiliation: European Lab Nonlinear Spect, Sesto Fiorentino, Italy; Univ Florence, Dept Phys & Astron, Sesto Fiorentino, Italy; Univ Florence, Dept Neurosci Psychol Drug Res & Child Hlth, Florence, Italy; CNR, Natl Inst Opt, Sesto Fiorentino, Italy; Univ Padua, Dept Biol, Padua, Italy; Univ Florence, Dept Biol, Sesto Fiorentino, Italy.

Abstract: Confocal detection in digital scanned laser light-sheet fluorescence microscopy (DSLM) has been established as a gold standard method to improve image quality. The selective line detection of a complementary metal-oxide-semiconductor camera (CMOS) working in rolling shutter mode allows the rejection of out-of-focus and scattered light, thus reducing background signal during image formation. Most modern CMOS have two rolling shutters, but usually only a single illuminating beam is used, halving the maximum obtainable frame rate. We report on the capability to recover the full image acquisition rate via dual confocal DSLM by using an acoustooptic deflector. Such a simple solution enables us to independently generate, control and synchronize two beams with the two rolling slits on the camera. We show that the doubling of the imaging speed does not affect the confocal detection high contrast. (C) The Authors. Published by SPIE under a Creative Commons Attribution 4.0 Unported License.

Journal/Review: JOURNAL OF BIOMEDICAL OPTICS

Volume: 24 (10)      Pages from: 106504-1  to: 106504-6

More Information: This project has received funding from the European Union´s Horizon 2020 Framework Programme for Research and Innovation under Grant Agreement No. 654148 (Laserlab-Europe), specific Grant Agreement No. 720270 (Human Brain Project SGA1), No. 785907 (Human Brain Project SGA2), and Marie Sklodowska-Curie Grant Agreement No. 793849 (MesoBrainMicr), from the General Hospital Corporation Center of the National Institutes of Health under award number 1U01MH117023-01, from the H2020 EXCELLENT SCIENCE-European Research Council (ERC) under Grant Agreement ID No. 692943 (BrainBIT) and from the Italian Ministry for Education in the framework of Eurobioimaging (ESFRI research infrastructure)-Advanced Light Microscopy Italian Node. The authors are grateful to Mr. Riccardo Ballerini from LENS mechanical workshop for technical assistance.
KeyWords: light-sheet microscopy; digital scanned laser light-sheet fluorescence microscopy; confocal detection; acousto-optic deflector; high-throughput microscopy; high contrast; mouse brain; zebrafish brain
DOI: 10.1117/1.JBO.24.10.106504

Citations: 23
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