In:
Frontiers in Bioengineering and Biotechnology, Frontiers Media SA, Vol. 8 ( 2021-1-7)
Abstract:
Minibrain is a 3D brain in vitro spheroid model, composed of a mixed population of neurons and glial cells, generated from human iPSC derived neural stem cells. Despite the advances in human 3D in vitro models such as aggregates, spheroids and organoids, there is a lack of labeling and imaging methodologies to characterize these models. In this study, we present a step-by-step methodology to generate human minibrain nurseries and novel strategies to subsequently label projection neurons, perform immunohistochemistry and 3D imaging of the minibrains at large multiplexable scales. To visualize projection neurons, we adapt viral transduction and to visualize the organization of cell types we implement immunohistochemistry. To facilitate 3D imaging of minibrains, we present here pipelines and accessories for one step mounting and clearing suitable for confocal microscopy. The pipelines are specifically designed in such a way that the assays can be multiplexed with ease for large-scale screenings using minibrains and other organoid models. Using the pipeline, we present (i) dendrite morphometric properties obtained from 3D neuron morphology reconstructions, (ii) diversity in neuron morphology, and (iii) quantified distribution of progenitors and POU3F2 positive neurons in human minibrains.
Type of Medium:
Online Resource
ISSN:
2296-4185
DOI:
10.3389/fbioe.2020.582650
DOI:
10.3389/fbioe.2020.582650.s001
DOI:
10.3389/fbioe.2020.582650.s002
DOI:
10.3389/fbioe.2020.582650.s003
DOI:
10.3389/fbioe.2020.582650.s004
DOI:
10.3389/fbioe.2020.582650.s005
DOI:
10.3389/fbioe.2020.582650.s006
DOI:
10.3389/fbioe.2020.582650.s007
DOI:
10.3389/fbioe.2020.582650.s008
DOI:
10.3389/fbioe.2020.582650.s009
DOI:
10.3389/fbioe.2020.582650.s010
DOI:
10.3389/fbioe.2020.582650.s011
DOI:
10.3389/fbioe.2020.582650.s012
DOI:
10.3389/fbioe.2020.582650.s013
DOI:
10.3389/fbioe.2020.582650.s014
DOI:
10.3389/fbioe.2020.582650.s015
DOI:
10.3389/fbioe.2020.582650.s016
Language:
Unknown
Publisher:
Frontiers Media SA
Publication Date:
2021
detail.hit.zdb_id:
2719493-0
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