CE42 - Capteurs, instrumentation

Super-resolution imaging through scattering media – SpeckleSTED

Submission summary

In recent years, super-resolution fluorescence microscopies have largely contributed to the understanding of many biological processes occurring on a nanometric scale in living matter. However, biological tissues are turbid. Beyond 100 microns in depth, focused coherent beams are then degraded into random spotty structures called “speckle patterns”. For this reason, imaging deep through tissues remains a challenge, seemingly particularly incompatible with super-resolution. Super-resolution in scattering samples remains mostly unexplored.

The SpeckleSTED project aims at exploring a novel approach to perform super-resolution imaging through scattering media, and in particular through tissues, by using speckle patterns and the properties of the diffusers to our advantage. Indeed, speckle patterns, which naturally appear in random media, also naturally contain the structures required to perform super-resolution microscopy.

The proposed technique is inspired by the recent demonstration that fluorescence imaging can be performed through strongly scattering media without any active control on the wavefront or any adaptive optical element. The image recorded with this technique is blurred due to the speckled intensity distribution of the excitation beam. However, it is possible to unblur the image thanks to iterative algorithms [1] recovering the object itself [2].

In the Neurophotonics Laboratory, I developed a super-resolution STED microscope, shortly following the invention of this technique that was awarded the Nobel Prize in chemistry in 2014. A STED microscope is basically a confocal microscope where a de-excitation laser beam is tailored with the shape of a donut centered on the diffraction-limited excitation spot. The resolution is improved by confining fluorescence to sub-diffraction dimensions, saturating the de-excitation probability with the donut beam. Breaking of the diffraction limit is obtained by saturating stimulated emission. The donut beam is typically engineered as an “optical vortex” beam: a beam with zero intensity along the optical axis and a helical phase around. The phase of the field is undefined (or “singular”) on this line.

Speckle patterns are usually described as spotty structures but they also contain a high density of optical vortices, such as the ones used for STED microscopy. Recent works demonstrated that speckle patterns could be used to improve the resolution of standard widefield microscopes even in the linear regime [3]. Here, I point out that the presence of optical vortices in speckle patterns make them ideal candidates as illumination patterns to perform super-resolution microscopy by saturating an optical transition. As a first step, we recently demonstrated [4] that optical vortices contained in random speckle patterns can confine fluorescence to sub-diffraction dimensions. This property so allowed us to achieve three-dimensional super-resolution speckle microscopy under saturated excitation conditions [5]. Moreover, since STED microscopy requires the use of two beams (excitation and de-excitation) exhibiting inverted intensity profiles, we also described a technique to exchange deterministically the location of vortices and maxima in a speckle pattern generated through an unknown diffuser [6].

SpeckleSTED thus aims at demonstrating super-resolution speckle imaging by stimulated emission through model diffusers and biological tissues.

1. Fienup, Appl. Opt., 1982. 21(15): p. 2758-2769.
2. Bertolotti, et al., Nature, 2012. 491(7423): p. 232-234.
3. Mudry, et al., Nat. Photon., 2012. 6(5): p. 312-315.
4. Pascucci, M., G. Tessier, V. Emiliani, and M. Guillon, Phys. Rev. Lett., 2016. 116(9): p. 093904.
5. Pascucci, M., Ganesan, S., Katz, O., Emiliani, V., Guillon, M., arXiv :1710.05056
6. Gateau, J., H. Rigneault, and M. Guillon, Phys. Rev. Lett. 2017. 118(4): p. 5.

Project coordinator

Monsieur Marc GUILLON (Neurophotonique)

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.


UPDescartes-UMR 8250 Neurophotonique

Help of the ANR 328,580 euros
Beginning and duration of the scientific project: December 2018 - 36 Months

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