Superresolved multiphoton microscopy with spatial frequency-modulated imaging

Superresolved far-field microscopy has emerged as a powerful tool for investigating the structure of objects with resolution well below the diffraction limit of light. Nearly all superresolution imaging techniques reported to date rely on real energy states of fluorescent molecules to circumvent the...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2016-06, Vol.113 (24), p.6605-6610
Main Authors: Field, Jeffrey J., Wernsing, Keith A., Domingue, Scott R., Motz, Alyssa M. Allende, DeLuca, Keith F., Levi, Dean H., DeLuca, Jennifer G., Young, Michael D., Squier, Jeff A., Bartels, Randy A.
Format: Article
Language:English
Subjects:
Atoms & subatomic particles
Diffraction
Fluorescence
harmonic generation
MATERIALS SCIENCE
Microscopy
Microscopy, Fluorescence, Multiphoton - methods
Models, Theoretical
multiphoton microscopy
Physical Sciences
Scattering
superresolution
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Summary:Superresolved far-field microscopy has emerged as a powerful tool for investigating the structure of objects with resolution well below the diffraction limit of light. Nearly all superresolution imaging techniques reported to date rely on real energy states of fluorescent molecules to circumvent the diffraction limit, preventing superresolved imaging with contrast mechanisms that occur via virtual energy states, including harmonic generation (HG). We report a superresolution technique based on spatial frequency-modulated imaging (SPIFI) that permits superresolved nonlinear microscopy with any contrast mechanism and with single-pixel detection. We show multimodal superresolved images with two-photon excited fluorescence (TPEF) and second-harmonic generation (SHG) from biological and inorganic media. Multiphoton SPIFI (MP-SPIFI) provides spatial resolution up to 2η below the diffraction limit, where η is the highest power of the nonlinear intensity response. MP-SPIFI can be used to provide enhanced resolution in optically thin media and may provide a solution for superresolved imaging deep in scattering media.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.1602811113