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Related Experiment Video

Updated: Jul 1, 2025

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Protocol to dissect and dissociate the mouse brainstem for single-cell RNA-seq applications.

Wiktor S Phillips1, Naify Ramadan1, Athina Samara2

  • 1Department of Women's and Children's Health, Karolinska Institutet, Stockholm, Sweden; Astrid Lindgren Children's Hospital, Karolinska University Hospital, Stockholm, Sweden.

STAR Protocols
|March 10, 2024
PubMed
Summary

This study presents a new protocol for processing neonatal mouse brainstem tissue for transcriptomics. The method enables asynchronous sample collection and cell processing, overcoming challenges with low cell yield from small brain structures.

Keywords:
Cell isolationDevelopmental biologyNeuroscienceSingle Cell

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Area of Science:

  • Neuroscience
  • Molecular Biology
  • Genomics

Background:

  • Transcriptomic analysis of small brain structures, such as brainstem nuclei, is hindered by low cell yields.
  • Efficient protocols are needed for dissecting, dissociating, and preserving these delicate tissues.

Purpose of the Study:

  • To develop and validate a protocol for processing neonatal mouse brainstem tissue for single-cell transcriptomics.
  • To enable asynchronous sample collection and downstream processing, improving cell yield and experimental flexibility.

Main Methods:

  • A protocol for dissection, dissociation, and cryopreservation of neonatal mouse brainstem tissue was established.
  • The method was demonstrated using the preBötzinger complex and SmartSeq3 cDNA library preparation.

Main Results:

  • The protocol successfully yields viable cells from small brainstem structures.
  • It allows for flexible, asynchronous sample collection and processing, addressing previous limitations.

Conclusions:

  • This cryopreservation and dissociation protocol is effective for neonatal mouse brainstem tissue.
  • The method is adaptable for various brainstem regions and transcriptomic library preparation techniques.