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Yide Zhang1,2, David Benirschke1, Ola Abdalsalam1

  • 1Department of Electrical Engineering, University of Notre Dame, Notre Dame, IN 46556, USA.

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This summary is machine-generated.

We introduce generalized stepwise optical saturation (GSOS), a novel super-resolution microscopy technique. GSOS extends stepwise optical saturation (SOS) and enables simultaneous super-resolution and fluorescence lifetime imaging microscopy (FLIM).

Keywords:
(100.6640) Superresolution(170.2520) Fluorescence microscopy(170.3650) Lifetime-based sensing(180.4315) Nonlinear microscopy(190.4180) Multiphoton processes

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Area of Science:

  • Optical Microscopy
  • Super-resolution Imaging
  • Biomedical Optics

Background:

  • Super-resolution microscopy enhances image detail beyond the diffraction limit.
  • Stepwise optical saturation (SOS) is a recently developed super-resolution technique.
  • Fluorescence lifetime imaging microscopy (FLIM) provides functional information about fluorophores.

Purpose of the Study:

  • To present and theoretically analyze a generalized stepwise optical saturation (GSOS) technique.
  • To demonstrate the super-resolution capabilities of GSOS.
  • To achieve simultaneous super-resolution and FLIM.

Main Methods:

  • Developed the theoretical basis of GSOS using a two-level fluorophore model and perturbation theory.
  • Performed theoretical analysis and numerical simulations for GSOS with sinusoidal and pulse-train modulations.
  • Experimentally implemented GSOS on a custom-built two-photon frequency-domain (FD) FLIM system.

Main Results:

  • GSOS generalizes and extends the SOS super-resolution microscopy technique.
  • GSOS increases imaging resolution by a factor of M.
  • Simultaneous super-resolution and FLIM were achieved using GSOS with pulse-train modulation.
  • Experimental demonstration of super-resolution FLIM on a cell sample.

Conclusions:

  • GSOS represents a significant advancement in super-resolution microscopy.
  • The technique enables simultaneous acquisition of high-resolution structural and functional information.
  • This work reports the first implementation of super-resolution imaging within frequency-domain FLIM.