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Tissue Handling and Dissociation for Single-Cell RNA-Seq.

Felipe A Vieira Braga1, Ricardo J Miragaia2

  • 1Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, UK. fvb@sanger.ac.uk.

Methods in Molecular Biology (Clifton, N.J.)
|April 28, 2019
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Summary

Obtaining high-quality human immune cells for single-cell analysis requires optimized tissue dissociation protocols. Minimizing stress signatures during isolation is crucial to prevent biased interpretation of single-cell data.

Keywords:
DigestionImmune cellsSingle-cell RNA sequencingSingle-cell suspensionTissue processing

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Area of Science:

  • Immunology
  • Cell Biology
  • Genomics

Background:

  • Single-cell analysis provides high-resolution insights into cellular heterogeneity.
  • Immune cells reside in diverse tissues, necessitating specific isolation techniques.
  • Tissue dissociation is a critical first step for single-cell studies.

Purpose of the Study:

  • To describe protocols for dissociating human lymphoid and nonlymphoid tissues for immune cell isolation.
  • To identify factors influencing single-cell data quality during tissue processing.
  • To highlight the impact of stress signatures on immune cell interpretation.

Main Methods:

  • Tissue dissociation techniques for lymphoid and nonlymphoid tissues.
  • Protocol optimization for obtaining viable human immune cells.
  • Analysis of stress signatures in isolated immune cells.

Main Results:

  • Established protocols for effective immune cell isolation from various human tissues.
  • Identified key factors, including stress signatures, affecting single-cell data integrity.
  • Demonstrated the potential for stress-induced artifacts in immune cell profiling.

Conclusions:

  • Optimized tissue dissociation protocols are essential for reliable single-cell immune cell studies.
  • Awareness and mitigation of stress signatures are critical for accurate data interpretation.
  • These protocols facilitate downstream single-cell analyses of human immune populations.