Modular multimodal platform for classical and high throughput light sheet microscopy

Abstract

Light-sheet fluorescence microscopy (LSFM) has become an important tool for biological and biomedical research. Although several illumination and detection strategies have been developed, the sample mounting still represents a cumbersome procedure as this is highly dependent on the type of sample and often this might be time consuming. This prevents the use of LSFM in other promising applications in which a fast and straightforward sample-mounting procedure and imaging are essential. These include the high-throughput research fields, e.g. in drug screenings and toxicology studies. Here we present a new imaging paradigm for LSFM, which exploits modularity to offer multimodal imaging and straightforward sample mounting strategy, enhancing the flexibility and throughput of the system. We describe its implementation in which the sample can be imaged either as in any classical configuration, as it flows through the light-sheet using a fluidic approach, or a combination of both. We also evaluate its ability to image a variety of samples, from zebrafish embryos and larvae to 3D complex cell cultures.

Multiple asters organize the yolk microtubule network during dclk2-GFP zebrafish epiboly

“…S1C). LSFM imaging was performed using a custom-made light-sheet set up 15 , called Flexi-SPIM. For illumination, we used 488, 561 and 637 nm lasers (Cobolt, MDL488, MLD561, MLD637) and two air objectives (Nikon 4 ×, NA 0.13).…”

“…Those observations have been quantified across many embryos thanks to the increased throughput capabilities of our custom-made LSFM microscope 15 . Our approach makes it possible to look at the embryo as a whole, to visualize the dynamic organization in 3D with high resolution (0.65 µm) and contrast, and to quantify the phenotypic variability that has remained unseen up to now.…”

“…The dclk2-GFP zebrafish transgenic line has been used as MT reporter line in several publications [14][15][16][17][18][19][20][21][22][23][24][25] to visualize MT dynamics in live cells. The Tg(ef1α:dclk-GFP) transgene contains two doublecortin domains (microtubule-binding domain) of zebrafish dclk1 fused to gfp, and under the control of the Xenopus ef1α promoter.…”

A minimal-complexity light-sheet microscope maps network activity in 3D neuronal systems

“…Functional activity mapping in various samples has been performed at scanning rates between 0.5 to 50 Hz (Suppl. Table 1) [31][32][33][34][35][36][37] . High-speed imaging using a single primary objective both for illumination and detection has become possible via oblique plane microscopy, introduced by C. Dunsby 38 .…”

A minimal-complexity light-sheet microscope maps network activity in 3D neuronal systems

“…Functional activity mapping in various samples has been performed at scanning rates between 0.5 and 50 Hz (Suppl. Table 1) [31][32][33][34][35][36][37] . High-speed imaging using a single primary objective both for illumination and detection has become possible via oblique plane microscopy, introduced by C. Dunsby 38 .…”