Oblique plane light-sheet microscopy provides versatile and rapid volumetric functional and structural fluorescence imaging for biology. A single microscope objective illuminates a tilted plane in the specimen and collects emission. Descanned emission is relayed and tilt-corrected through two additional microscope arms to a camera. Due to relative system complexity, estimating required hardware for diverse applications and system alignment can be challenging. Towards overcoming this obstacle and maximizing access, we present an open access scanned oblique plane microscopy platform, along with a design GUI, detailed alignment protocols, control software, application examples in zebrafish and mice, as well as possible systems extensions.
Light-sheet microscopy has evolved as an indispensable tool in imaging biological samples. It can image 3D samples at fast speed, with high-resolution optical sectioning, and with reduced photobleaching effects. These properties make light-sheet microscopy ideal for imaging fluorophores in a variety of biological samples and organisms, e.g. zebrafish, drosophila, cleared mouse brains, etc. While most commercial turnkey light-sheet systems are expensive, the existing lower cost implementations, e.g. OpenSPIM, are focused on achieving high-resolution imaging of small samples or organisms like zebrafish. In this work, we substantially reduce the cost of light-sheet microscope system while targeting to image much larger samples, i.e. cleared mouse brains, at single-cell resolution. The expensive components of a lightsheet system – excitation laser, water-immersion objectives, and translation stage – are replaced with an incoherent laser diode, dry objectives, and a custom-built Arduino-controlled translation stage. A low-cost CUBIC protocol is used to clear fixed mouse brain samples. The open-source platforms of μManager and Fiji support image acquisition, processing, and visualization. Our system can easily be extended to multi-color light-sheet microscopy.
The ability to image through a scattering or diffusive medium such as tissue or hazy atmosphere is a goal which has
garnered extensive attention from the scientific community. Existing imaging methods in this field make use of phase
conjugation, time of flight, iterative wave-front shaping or statistical averaging approaches, which tend to be either time
consuming or complicated to implement. We introduce a novel and practical way of statistical averaging which makes
use of a rotating ground glass diffuser to nullify the adverse effects caused by speckle introduced by a first static diffuser
/ aberrator. This is a Fourier transform-based, holographic approach which demonstrates the ability to recover detailed
images and shows promise for further remarkable improvement. The present experiments were performed with 2D flat
images, but this method could be easily adapted for recovery of 3D extended object information. The simplicity of the
approach makes it fast, reliable, and potentially scalable as a portable technology. Since imaging through a diffuser has
direct applications in biomedicine and defense technologies this method may augment advanced imaging capabilities in
many fields.
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