Preparation of the intact rodent organ of Corti for RNAscope and immunolabeling, confocal microscopy, and quantitative analysis

Daniel O J Reijntjes; J Lukas Breitzler; Dora Persic; Sonja J Pyott

doi:10.1016/j.xpro.2021.100544

Preparation of the intact rodent organ of Corti for RNAscope and immunolabeling, confocal microscopy, and quantitative analysis

STAR Protoc. 2021 May 24;2(2):100544. doi: 10.1016/j.xpro.2021.100544. eCollection 2021 Jun 18.

Authors

Daniel O J Reijntjes¹, J Lukas Breitzler², Dora Persic², Sonja J Pyott²

Affiliations

¹ The Center for Hearing and Balance, Department of Otolaryngology-Head and Neck Surgery, The Johns Hopkins University School of Medicine, Baltimore, MA 21201, USA.
² Department of Otorhinolaryngology-Head and Neck Surgery, University Medical Center Groningen, University of Groningen, 9713GZ Groningen, the Netherlands.

Abstract

This protocol describes the preparation of the mouse organ of Corti for RNAscope, immunolabeling, confocal microscopy, and quantitative image analysis to examine transcript and protein localization, sensory hair cells, and synapses. This protocol can be applied to mice and other rodents (juvenile and adult) and can be adapted for other techniques, including electrophysiology and RNA sequencing. This protocol features minimal tissue processing to preserve viability for downstream assays, while isolating the organ of Corti is the most challenging step. For additional details on the use and execution of this protocol, please refer to McLean et al. (2009); Schuth et al. (2014); Lingle et al. (2019); Pyott et al. (2020).

Keywords: Antibody; In Situ Hybridization; Microscopy; Molecular/Chemical Probes; Neuroscience.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Immunohistochemistry / methods*
In Situ Hybridization
Mice
Microscopy, Confocal / methods*
Organ of Corti* / chemistry
Organ of Corti* / physiology
RNA / analysis*
Single Molecule Imaging / methods*

Substances

RNA