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. 2020 May 12;13:75. doi: 10.3389/fnmol.2020.00075

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Copyright © 2020 Tsuneoka and Funato.

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) and the copyright owner(s) 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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Detection of low abundance mRNA using in situ HCR. (A,D) In situ HCR using ATTO550-conjugated hairpin DNA (#S23) detected Oxtr mRNA in mouse brain. Amplification was performed overnight. (B,E) Fluorescent ISH with tyramide signal amplification for Oxtr mRNA. Oxtr mRNA was visualized by a combination of DIG-labeled probe, peroxidase-conjugated anti-DIG antibody, and tyramide Alexa568. (C,F) Chromogenic ISH for Oxtr mRNA by a combination of DIG-labeled probe, alkaline phosphatase-conjugated anti-DIG antibody, and BCIP/NBT. Color development was performed for 3 days. Panels (D–F) corresponds to dashed squares in panels (A–C), respectively. Panels (G–I) corresponds to dashed squares in (D–F). Scale bars: 400 μm (A–C), 100 μm (D–F) and 10 μm (G–I). BNST, bed nucleus of stria terminalis; BNSTpr, principal nucleus of BNST; BNSTrh, rhomboid nucleus of BNST; LPOA, lateral preoptic area; MN, magnocellular nucleus; MPOA, medial preoptic area.