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  3. Protocol for in vivo elimination of avian auditory hair cells, multiplexed mRNA detection, immunohistochemistry, and S-phase labeling

Protocol for in vivo elimination of avian auditory hair cells, multiplexed mRNA detection, immunohistochemistry, and S-phase labeling

  • STAR Protoc. 2024 Jun 21;5(2):103118. doi: 10.1016/j.xpro.2024.103118.
Mitsuo P Sato 1,  Austin P Huang 2,  Stefan Heller 2,  Nesrine Benkafadar 3
Affiliations

Affiliations

  • 1 Department of Otolaryngology - Head and Neck Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Otolaryngology-Head and Neck Surgery, Kindai University School of Medicine, Osaka, Japan.
  • 2 Department of Otolaryngology - Head and Neck Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
  • 3 Department of Otolaryngology - Head and Neck Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA. Electronic address: kafadar@stanford.edu.
PMID: 38852155 DOI: 10.1016/j.xpro.2024.103118
Abstract

The avian inner ear can naturally regenerate sensory hair cells and is therefore an ideal candidate for investigating mechanisms leading to hair cell regeneration and functional recovery. Here, we present a surgical protocol for eliminating auditory hair cells via sisomicin injection into the lateral semicircular canal. We describe steps for multiplex mRNA detection in chicken basilar papilla and utricle sections. We then detail procedures for integrating immunohistochemistry for concurrent mRNA and protein visualization, complemented by S-phase labeling with EdU. For complete details on the use and execution of this protocol, please refer to Benkafadar et al., Benkafadar et al., Sato et al., Janesick et al., Scheibinger et al.1,2,3,4,5.

Keywords

antibody; in situ hybridization; model organisms.

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