Dual-color on-chip light sheet microscopy of drosophila embryos

摘要

Drosophila Melanogaster is a sample of high biological interest that is being widely used as biological model, due tothe relatively short life cycle, short genome and ease in culturing. In this work we present a microscope on chipcapable of processing Drosophila embryos to obtain three dimensional fluorescent images at high throughput. Thisdevice, based on light sheet microscopy, uses a plane of light intercepting the sample channel to optically and noninvasivelysection the embryos while flowing. This permits to automatically acquire for each sample the stack ofimages necessary for the subsequent 3D reconstruction with no need of any manual sample positioning andalignment. The whole chip is fabricated in a glass substrate by femtosecond laser micromachining. The device hasbeen optimized for the specific morphology of the sample. Indeed, the highly elliptical shape of the embryos (about100 x 500 μm2) might affect the image quality degrading both the vertical and the axial resolution of the system. Toovercome this issue, we have first optimized the layout of the fluidic channel to precisely control the sampleorientation by means of hydrodynamic forces. Thereafter, we have optimized the properties of the optical circuit, torealize two opposite light sheets impinging on the sample, perfectly overlapped, with a high signal to noise ratio. Withthese actions, we have been able to obtain high quality Drosophila reconstruction.