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Flow-cytometry-based protocols for human blood/marrow immunophenotyping with minimal sample perturbation.

Laura G Rico1, Roser Salvia1, Michael D Ward2

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STAR Protocols
|October 25, 2021
PubMed
Summary

This protocol enhances flow cytometry by skipping harsh sample preparation steps. This allows for better analysis of live cells and identification of rare cell populations.

Keywords:
CancerClinical ProtocolFlow Cytometry/Mass CytometryHealth SciencesImmunologyStem Cells

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Area of Science:

  • Immunology
  • Cell Biology
  • Biotechnology

Background:

  • Traditional flow cytometry sample preparation involves steps like erythrolysis, density gradients, and washing.
  • These steps can cause cell loss and alter cell surface antigen expression, potentially affecting analysis accuracy.
  • Minimizing sample perturbation is crucial for studying live cells and identifying rare populations.

Purpose of the Study:

  • To present a protocol for improved flow cytometry analysis of marrow and peripheral blood cells.
  • To introduce two basic approaches for identifying cell surface antigens with minimal sample perturbation.
  • To highlight the advantages of reduced sample preparation for live cell analysis.

Main Methods:

  • The protocol avoids erythrolytic solutions, density gradients, and washing steps.
  • Two distinct approaches are described for identifying cell surface antigens.
  • The methods focus on preserving sample integrity and minimizing cellular stress.

Main Results:

  • The described approaches have been successfully used to identify both healthy and pathologically rare cells.
  • Minimal sample perturbation leads to improved preservation of live cells.
  • The protocol allows for more accurate study of cell surface antigens.

Conclusions:

  • This protocol offers a simplified and effective method for flow cytometry analysis.
  • Avoiding harsh preparation steps enhances the study of live cells and rare cell populations.
  • The presented techniques minimize unwanted effects from sample preparation, improving analytical outcomes.