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. 2018 Jan 17;11:750. doi: 10.3389/fnins.2017.00750

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Copyright © 2018 Lindsey, Douek, Loosli and Kaslin.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Compatibility of immunohistochemistry with OPT pipeline. (A) Successful double-labeling and OPT scanning of EdU (green) and the glial marker, glutamine synthetase (GS; red) in the adult zebrafish forebrain. White box denotes images shown in (B,C). (B,C) Co-labeling with DAPI (blue; B) and single (C) antibody labeling of GS (red) in cryosectioned, confocal-imaged tissue confirming the specificity of GS labeling shown using our OPT pipeline. (D,E) Whole brain immunohistochemistry in the adult zebrafish using a rabbit-anti-GFP primary antibody conjugated to Alexa 555 (D) recapitulates the same staining pattern seen with the endogenous GFP reporter in the Tg(Her4.1:gfp) transgenic line (E) prior to dehydration and clearing. White boxes in (D,E) depicts location of image displayed in (F). (F) GFP-positive staining in the deep quiescent glial layer of the periventricular gray zone of the adult optic tectum shown in cryosectioned, confocal-imaged tissue in the Tg(Her4.1:gfp) line mimicking the GFP pattern seen in tissue prepared using our OPT pipeline. (G–I) Examples of poor antibody penetration (G), poor immuno-labeling, and (I) quenched antibody staining post-OPT imaging using different GFP antibodies. (J–L) Adult control brain (J) in the Tg(mpeg1:gfp) macrophage line injected with EdU compared with brains post telencephalic lesion (yellow symbol) examined at 1-day post lesion (dpl; K) and 7-dpl (L) for changes in macrophage distribution (purple) and cell proliferation (green). Scale bars: A, G = 300 μm; B-C = 150 μm; F = 200 μm; J-K = 500 μm.