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Updated: May 4, 2026

Internalization and Observation of Fluorescent Biomolecules in Living Microorganisms via Electroporation
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An Electroporation Cytometry Protocol for Live-Cell, Fluorescence Microscopy Using U2 OS Cell Culture.

Thomas Nesmith1, Christian Vieira2, Darius G Rackus1

  • 1Department of Chemistry and Biology, Toronto Metropolitan University; Institute for Biomedical Engineering, Science and Technology (iBEST), a partnership between St. Michael's Hospital, a site of Unity Health Toronto, and Toronto Metropolitan University; Keenan Research Centre for Biomedical Science at St. Michael's Hospital.

Journal of Visualized Experiments : Jove
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PubMed
Summary
This summary is machine-generated.

Pulsed electric fields (PEFs) enable gene electrotransfer and cancer treatment. A new system allows live-cell microscopy to track cell cycle changes after PEF exposure, improving electroporation techniques.

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Area of Science:

  • Biomedical Engineering
  • Cell Biology
  • Medical Physics

Background:

  • Pulsed electric fields (PEFs) are crucial in medical research, particularly for electroporation techniques like gene electrotransfer (GET) and electrochemotherapy (ECT).
  • While effective, current electroporation methods require advanced supporting technologies for detailed live-cell analysis and improved efficacy.

Purpose of the Study:

  • To develop an advanced electroporation cytometry system for live-cell experimentation with long-term fluorescence microscopy.
  • To assess real-time cell cycle changes in response to PEF exposure using a FUCCI(CA)5 U2-OS cell line.

Main Methods:

  • Development of an electroporation cytometry system integrated with long-term fluorescence microscopy capabilities.
  • Utilized a stably expressing FUCCI(CA)5 U2-OS cell line to monitor cell cycle dynamics.
  • Conducted timelapse microscopy for up to 30 hours to observe cellular responses to PEF exposure.

Main Results:

  • Achieved up to 30 hours of timelapse microscopy, enabling real-time observation of cell activity.
  • Quantified phase-specific changes in the cell cycle in response to PEF treatment.
  • Demonstrated the system's capability to support detailed live-cell PEF experimentation.

Conclusions:

  • The developed electroporation cytometry system effectively supports live-cell PEF experimentation and long-term monitoring.
  • This technology provides a valuable tool for quantifying PEF-induced cell cycle alterations.
  • The findings contribute to the advancement of electroporation techniques in biomedical research.