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Enhancement of image quality and imaging depth with Airy light-sheet microscopy in cleared and non-cleared neural tissue.

ABSTRACT: We have investigated the effect of Airy illumination on the image quality and depth penetration of digitally scanned light-sheet microscopy in turbid neural tissue. We used Fourier analysis of images acquired using Gaussian and Airy light-sheets to assess their respective image quality versus penetration into the tissue. We observed a three-fold average improvement in image quality at 50 ?m depth with the Airy light-sheet. We also used optical clearing to tune the scattering properties of the tissue and found that the improvement when using an Airy light-sheet is greater in the presence of stronger sample-induced aberrations. Finally, we used homogeneous resolution probes in these tissues to quantify absolute depth penetration in cleared samples with each beam type. The Airy light-sheet method extended depth penetration by 30% compared to a Gaussian light-sheet.

SUBMITTER: Nylk J

PROVIDER: S-EPMC5102539 | biostudies-literature | 2016 Oct

REPOSITORIES: biostudies-literature

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Enhancement of image quality and imaging depth with Airy light-sheet microscopy in cleared and non-cleared neural tissue.

Nylk Jonathan J McCluskey Kaley K Aggarwal Sanya S Tello Javier A JA Dholakia Kishan K

Biomedical optics express 20160914 10

We have investigated the effect of Airy illumination on the image quality and depth penetration of digitally scanned light-sheet microscopy in turbid neural tissue. We used Fourier analysis of images acquired using Gaussian and Airy light-sheets to assess their respective image quality versus penetration into the tissue. We observed a three-fold average improvement in image quality at 50 μm depth with the Airy light-sheet. We also used optical clearing to tune the scattering properties of the ti ...[more]

PMID: 27867712

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Project description:BackgroundThe inability to observe relevant biological processes in vivo significantly restricts human neurodevelopmental research. Advances in appropriate in vitro model systems, including patient-specific human brain organoids and human cortical spheroids (hCSs), offer a pragmatic solution to this issue. In particular, hCSs are an accessible method for generating homogenous organoids of dorsal telencephalic fate, which recapitulate key aspects of human corticogenesis, including the formation of neural rosettes-in vitro correlates of the neural tube. These neurogenic niches give rise to neural progenitors that subsequently differentiate into neurons. Studies differentiating induced pluripotent stem cells (hiPSCs) in 2D have linked atypical formation of neural rosettes with neurodevelopmental disorders such as autism spectrum conditions. Thus far, however, conventional methods of tissue preparation in this field limit the ability to image these structures in three-dimensions within intact hCS or other 3D preparations. To overcome this limitation, we have sought to optimise a methodological approach to process hCSs to maximise the utility of a novel Airy-beam light sheet microscope (ALSM) to acquire high resolution volumetric images of internal structures within hCS representative of early developmental time points.ResultsConventional approaches to imaging hCS by confocal microscopy were limited in their ability to image effectively into intact spheroids. Conversely, volumetric acquisition by ALSM offered superior imaging through intact, non-clarified, in vitro tissues, in both speed and resolution when compared to conventional confocal imaging systems. Furthermore, optimised immunohistochemistry and optical clearing of hCSs afforded improved imaging at depth. This permitted visualization of the morphology of the inner lumen of neural rosettes.ConclusionWe present an optimized methodology that takes advantage of an ALSM system that can rapidly image intact 3D brain organoids at high resolution while retaining a large field of view. This imaging modality can be applied to both non-cleared and cleared in vitro human brain spheroids derived from hiPSCs for precise examination of their internal 3D structures. This process represents a rapid, highly efficient method to examine and quantify in 3D the formation of key structures required for the coordination of neurodevelopmental processes in both health and disease states. We posit that this approach would facilitate investigation of human neurodevelopmental processes in vitro.

Dataset Information

Enhancement of image quality and imaging depth with Airy light-sheet microscopy in cleared and non-cleared neural tissue.

Publications

Enhancement of image quality and imaging depth with Airy light-sheet microscopy in cleared and non-cleared neural tissue.

Similar Datasets

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