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Related Experiment Video

Updated: Apr 23, 2026

Multiplexed Barcoding Image Analysis for Immunoprofiling and Spatial Mapping Characterization in the Single-Cell Analysis of Paraffin Tissue Samples
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Transient partial permeabilization with saponin enables cellular barcoding prior to surface marker staining.

Gregory K Behbehani1, Colin Thom, Eli R Zunder

  • 1Baxter Laboratory for Stem Cell Biology, Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California; Divisions of Hematology and Oncology, Department of Medicine, Stanford University School of Medicine, Stanford, California.

Cytometry. Part a : the Journal of the International Society for Analytical Cytology
|October 3, 2014
PubMed
Summary

This study introduces a new method for cellular barcoding that preserves surface marker staining. This technique enhances high-throughput cytometric analysis by simplifying protocols and improving data reliability.

Keywords:
fluorescent cellular barcodingmass cytometrymass-tag cellular barcodingpermeabilizationsaponin

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Area of Science:

  • Immunology
  • Cell Biology
  • Biotechnology

Background:

  • Fluorescent cellular barcoding and mass-tag cellular barcoding are advanced cytometric techniques.
  • These methods increase sample throughput, reduce inter-sample variation, and conserve reagents.
  • Existing protocols require barcoding post-staining, hindering analysis of alcohol-sensitive epitopes.

Purpose of the Study:

  • To develop an improved cellular barcoding protocol.
  • To enable barcoding of fixed cells while preserving surface marker staining.
  • To simplify barcoding procedures for cytometric analysis.

Main Methods:

  • A novel method involving transient partial permeabilization with 0.02% saponin was employed.
  • Fixed cells were barcoded using fluorescent or mass-tagged reagents after partial permeabilization.
  • The protocol was designed to maintain the integrity of surface marker staining.

Main Results:

  • The new method achieved efficient and consistent barcode staining.
  • Surface marker staining integrity was successfully preserved.
  • The protocol simplifies barcoding and allows for reliable comparison of surface marker expression across samples.

Conclusions:

  • This optimized barcoding protocol overcomes limitations of previous methods.
  • It facilitates the reliable detection of subtle differences in surface marker expression.
  • The technique enhances the utility of cellular barcoding in cytometric applications.