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Development of a robust immuno-S-FISH protocol using imaging flow cytometry.

Kathryn A Fuller1, Sophia Bennett2, Henry Hui1

  • 1Translational Cancer Pathology Laboratory, School of Pathology and Laboratory Medicine, The University of Western Australia, Crawley, Australia.

Cytometry. Part a : the Journal of the International Society for Analytical Cytology
|May 5, 2016
PubMed
Summary

A novel immuno-S-FISH method combines immunophenotyping and fluorescence in situ hybridization (FISH) for cells in suspension. This technique enables automated, quantitative analysis of large cell populations for chromosomal abnormalities and cell phenotype.

Keywords:
S-FISHhematological malignancyimaging cytometryimmunophenotyping

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Area of Science:

  • Genetics
  • Cell Biology
  • Biotechnology

Background:

  • Fluorescence in situ hybridization (FISH) is a microscopy technique for detecting DNA sequences in cells.
  • Standard FISH typically analyzes cells on slides, limiting throughput and requiring specific sample preparation.
  • Immuno-FISH and suspension FISH (S-FISH) are modifications offering enhanced detection capabilities.

Purpose of the Study:

  • To develop and optimize a novel immuno-S-FISH method.
  • To combine immunophenotyping with FISH analysis in a single platform.
  • To enable high-throughput, quantitative analysis of cells in suspension.

Main Methods:

  • Development of an imaging immuno-S-FISH protocol.
  • Utilizing an imaging flow cytometer for simultaneous microscopy and flow cytometry.
  • Automated analysis of cell phenotype and FISH probe "spot" detection in suspension.

Main Results:

  • Successful integration of immunophenotyping and FISH on cells in suspension.
  • Demonstration of automated, quantitative "spot" counting and cell population analysis.
  • Optimization of the imaging immuno-S-FISH flow cytometry protocol for robustness.

Conclusions:

  • The developed immuno-S-FISH method offers a robust, single-platform solution for analyzing large cell numbers.
  • This technique enables simultaneous assessment of cell phenotype and chromosomal determinants.
  • The method has significant potential for various clinical applications requiring quantitative cell analysis.