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Updated: May 19, 2026
Triggering Cell Stress and Death Using Conventional UV Laser Confocal Microscopy
Published on: February 3, 2017
Singlet-oxygen-mediated cell death using spatially-localized two-photon excitation of an extracellular sensitizer
Frederico M Pimenta1, Rasmus L Jensen, Lotte Holmegaard
1Center for Oxygen Microscopy and Imaging, Department of Chemistry, Aarhus University , Aarhus 8000, Denmark.
Abstract:
Controlling and quantifying the photosensitized production of singlet oxygen are key aspects in mechanistic studies of oxygen-dependent photoinitiated cell death. In this regard, the commonly accepted practice of using intracellular photosensitizers is, unfortunately, plagued by problems that include the inability to accurately (1) quantify the sensitizer concentration in the irradiated domain and (2) control the local environment that influences light delivery and sensitizer photophysics. However, capitalizing on the fact that singlet oxygen produced outside a cell is also cytotoxic, many of these problems can be avoided with the use of an extracellular sensitizer. For the present study, a hydrophilic dendrimer-encased membrane-impermeable sensitizer was used to generate an extracellular population of singlet oxygen upon spatially localized two-photon irradiation. Through the use of this sensitizer and this approach, it is now possible to better control the singlet oxygen dose in microscope-based time- and space-resolved single cell experiments. Thus, we provide a solution to a limiting problem in mechanistic studies of singlet-oxygen-mediated cell death.
Insights
Researchers developed a new method to control singlet oxygen production outside cells, improving studies on cell death. This extracellular approach overcomes limitations of traditional intracellular photosensitizers for accurate mechanistic investigations.
Area of Science:
- Biochemistry
- Cell Biology
- Photochemistry
Background:
- Accurate control and quantification of singlet oxygen are crucial for studying photoinitiated cell death.
- Intracellular photosensitizers present challenges in quantifying sensitizer concentration and controlling the local cellular environment.
Purpose of the Study:
- To develop a method for controlling extracellular singlet oxygen production for mechanistic studies of cell death.
- To overcome the limitations associated with intracellular photosensitizers.
Main Methods:
- Utilized a hydrophilic dendrimer-encased, membrane-impermeable photosensitizer.
- Generated extracellular singlet oxygen via spatially localized two-photon irradiation.
- Employed a microscope-based setup for time- and space-resolved single-cell experiments.
Main Results:
- Successfully generated extracellular singlet oxygen using the novel photosensitizer system.
- Demonstrated improved control over singlet oxygen dose in cellular experiments.
- Overcame limitations in quantifying sensitizer concentration and controlling the local environment.
Conclusions:
- The extracellular photosensitizer approach offers a viable solution for mechanistic studies of singlet oxygen-mediated cell death.
- This method enhances the precision and reliability of experiments investigating oxygen-dependent cell death pathways.
- Provides a new tool for precise control of singlet oxygen generation in cellular research.
