ArticleBiomedical optics express2016
Wide-field medium-repetition-rate multiphoton microscopy reduces photodamage of living cells.
Article in Biomedical optics express, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 12 papers.
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Who cites it
12 citing papers in PubMed.
- Evaluation of a nanosecond dissipative soliton resonance fiber laser for multiphoton microscopy.Scientific reports · 2025Article
- SNR enhanced high-speed two-photon microscopy using a pulse picker and time gating detection.Scientific reports · 2023Article
- Label-free photothermal optical coherence microscopy to locate desired regions of interest in multiphoton imaging of volumetric specimens.Scientific reports · 2023Article
- Second harmonic generation microscopy: a powerful tool for bio-imaging.Biophysical reviews · 2023Review
- Photoresponsive Nanocarriers Based on Lithium Niobate Nanoparticles for Harmonic Imaging and On-Demand Release of Anticancer Chemotherapeutics.ACS nanoscience Au · 2022Article
- Phototoxicity induced in living HeLa cells by focused femtosecond laser pulses: a data-driven approach.Biomedical optics express · 2021Article
- Infrared Excitation Induces Heating and Calcium Microdomain Hyperactivity in Cortical Astrocytes.Biophysical journal · 2020Article
- Multiphoton microscopy: a personal historical review, with some future predictions.Journal of biomedical optics · 2020Article
- Mapping of real-time morphological changes in the neuronal cytoskeleton with label-free wide-field second-harmonic imaging: a case study of nocodazole.Neurophotonics · 2019Article
- Live imaging of contracting muscles with wide-field second harmonic generation microscopy using a high power laser.Biomedical optics express · 2019Article
- Optical Tweezers: Phototoxicity and Thermal Stress in Cells and Biomolecules.Micromachines · 2019Review
- Membrane water for probing neuronal membrane potentials and ionic fluxes at the single cell level.Nature communications · 2018Article
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Authors and funding
4 authors.
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Abstract
Demands of higher spatial and temporal resolutions in linear and nonlinear imaging keep pushing the limits of optical microscopy. We showed recently that a multiphoton microscope with 200 kHz repetition rate and wide-field illumination has a 2-3 orders of magnitude improved throughput compared to a high repetition rate confocal scanning microscope. Here, we examine the photodamage mechanisms and thresholds in live cell imaging for both systems. We first analyze theoretically the temperature increase in an aqueous solution resulting from illuminating with different repetition rates (keeping the deposited energy and irradiated volume constant). The analysis is complemented with photobleaching experiments of a phenolsulfonphthalein (phenol red) solution. Combining medium repetition rates and wide-field illumination promotes thermal diffusivity, which leads to lower photodamage and allows for higher peak intensities. A three day proliferation assay is also performed on living cells to confirm these results: dwell times can be increased by a factor of 3×10(6) while still preserving cell proliferation. By comparing the proliferation data with the endogenous two-photon fluorescence decay, we propose to use the percentage of the remaining endogenous two-photon fluorescence after exposure as a simple in-situ viability test. These findings enable the possibility of long-term imaging and reduced photodamage.
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