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Updated: Mar 25, 2026

Simultaneous Multicolor Imaging of Biological Structures with Fluorescence Photoactivation Localization Microscopy

Simultaneous Multicolor Imaging of Biological Structures with Fluorescence Photoactivation Localization Microscopy

Published on: December 9, 2013

Multicolor multiphoton microscopy based on a nanosecond supercontinuum laser source.

Claire Lefort¹, Rodney P O'Connor², Véronique Blanquet²

¹Université de Limoges, CNRS UMR 7252, Labex "Sigma-Lim", INRA UMR 1061, F-87000, Limoges, France. claire.lefort@xlim.fr.

Journal of Biophotonics

|February 13, 2016

Summary

This summary is machine-generated.

This study demonstrates multicolor multiphoton microscopy using a novel, wide spectral bandwidth excitation source. This breakthrough enables detailed in vivo imaging of biological samples, advancing nonlinear imaging capabilities.

Keywords:

multicolor imaging multiphoton microscopy nanosecond supercontinuum source nonlinear imaging

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Biomolecular Imaging of Cellular Uptake of Nanoparticles using Multimodal Nonlinear Optical Microscopy

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Published on: May 16, 2022

Area of Science:

Nonlinear Optics
Biomedical Imaging
Microscopy

Background:

Multiphoton microscopy offers intrinsic optical sectioning and deep tissue penetration.
Previous multiphoton microscopy systems were limited by narrow excitation spectral bandwidths.
Achieving multicolor imaging often requires multiple lasers or complex setups.

Purpose of the Study:

To demonstrate multicolor multiphoton microscopy with an unprecedented 300 nm excitation spectral bandwidth.
To showcase the utility of this wide bandwidth for in vivo biological imaging.
To compare image quality with conventional multiphoton microscopy techniques.

Main Methods:

Implementation of a compact, simple nanosecond supercontinuum (SC) source with a low repetition rate.
Excitation of samples over a 300 nm spectral bandwidth (full width half maximum).
Acquisition of multicolor images from fluorescently labeled biological samples.

Main Results:

Successful in vivo multicolor imaging of glioma tumor cells (eGFP) and mouse brain vasculature (Texas Red).
Demonstration that the wide spectral bandwidth covers the emission spectra of both tested fluorophores.
Comparable image quality achieved in vitro using the nanosecond SC source versus a conventional femtosecond laser.

Conclusions:

The developed nanosecond supercontinuum source enables a novel, wide-bandwidth approach to multiphoton microscopy.
This technology facilitates comprehensive biological characterization and advanced contrast mechanisms in nonlinear imaging.
Opens possibilities for multi-color imaging, diverse fluorophore utilization, and other multiphoton processes like three-photon microscopy.