title: Development of a real-time flexible multiphoton microendoscope for label-free imaging in a live animal.
abstract: We present a two-photon microendoscope capable of in vivo label-free deep-tissue high-resolution fast imaging through a very long optical fiber. First, an advanced light-pulse spectro-temporal shaping device optimally precompensates for linear and nonlinear distortions occurring during propagation within the endoscopic fiber. This enables the delivery of sub-40-fs duration infrared excitation pulses at the output of 5 meters of fiber. Second, the endoscopic fiber is a custom-made double-clad polarization-maintaining photonic crystal fiber specifically designed to optimize the imaging resolution and the intrinsic luminescence backward collection. Third, a miniaturized fiber-scanner of 2.2 mm outer diameter allows simultaneous second harmonic generation (SHG) and two-photon excited autofluorescence (TPEF) imaging at 8 frames per second. This microendoscope's transverse and axial resolutions amount respectively to 0.8 μm and 12 μm, with a field-of-view as large as 450 μm. This microendoscope's unprecedented capabilities are validated during label-free imaging, ex vivo on various fixed human tissue samples, and in vivo on an anesthetized mouse kidney demonstrating an imaging penetration depth greater than 300 μm below the surface of the organ. The results reported in this manuscript confirm that nonlinear microendoscopy can become a valuable clinical tool for real-time in situ assessment of pathological states.
authors.affiliation: XLIM, UMR-CNRS 7252, Université de Limoges, France.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Ducourthial
authors.affiliation: XLIM, UMR-CNRS 7252, Université de Limoges, France.
authors.forename: Pierre
authors.initials: P
authors.lastname: Leclerc
authors.affiliation: XLIM, UMR-CNRS 7252, Université de Limoges, France.
authors.forename: Tigran
authors.initials: T
authors.lastname: Mansuryan
authors.affiliation: XLIM, UMR-CNRS 7252, Université de Limoges, France.
authors.forename: Marc
authors.initials: M
authors.lastname: Fabert
authors.affiliation: XLIM, UMR-CNRS 7252, Université de Limoges, France.
authors.forename: Julien
authors.initials: J
authors.lastname: Brevier
authors.affiliation: PhLAM, UMR-CNRS 8523, Université Lille I, Villeneuve d'Ascq, France.
authors.forename: Rémi
authors.initials: R
authors.lastname: Habert
authors.affiliation: PhLAM, UMR-CNRS 8523, Université Lille I, Villeneuve d'Ascq, France.
authors.forename: Flavie
authors.initials: F
authors.lastname: Braud
authors.affiliation: CRIBL, UMR-CNRS 7276, Université de Limoges, France.
authors.forename: Renaud
authors.initials: R
authors.lastname: Batrin
authors.affiliation: Université Pierre et Marie Curie-Paris 06, LJP, F-75005 Paris, France.|CNRS, UMR 8237, LJP, F-75005 Paris, France.
authors.forename: Christine
authors.initials: C
authors.lastname: Vever-Bizet
authors.affiliation: Université Pierre et Marie Curie-Paris 06, LJP, F-75005 Paris, France.|CNRS, UMR 8237, LJP, F-75005 Paris, France.
authors.forename: Geneviève
authors.initials: G
authors.lastname: Bourg-Heckly
authors.affiliation: Laboratoire LITIS-QuantIF, EA 4108, Clinique Pneumologique, CHU de Rouen, France.
authors.forename: Luc
authors.initials: L
authors.lastname: Thiberville
authors.affiliation: CRIBL, UMR-CNRS 7276, Université de Limoges, France.
authors.forename: Anne
authors.initials: A
authors.lastname: Druilhe
authors.affiliation: PhLAM, UMR-CNRS 8523, Université Lille I, Villeneuve d'Ascq, France.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Kudlinski
authors.affiliation: XLIM, UMR-CNRS 7252, Université de Limoges, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Louradour
mesh_terms: D000818:Animals; D003952:Diagnostic Imaging / Q000295:instrumentation / Q000379:methods*; D004724:Endoscopy / Q000295:instrumentation / Q000379:methods*; D005355:Fibrosis / Q000473:pathology; D006801:Humans; D007668:Kidney / Q000033:anatomy & histology*; D007674:Kidney Diseases / Q000473:pathology*; D008168:Lung / Q000033:anatomy & histology; D051379:Mice; D036641:Microscopy, Fluorescence, Multiphoton / Q000295:instrumentation / Q000379:methods*; D017711:Nonlinear Dynamics; D055100:Optical Fibers; D015203:Reproducibility of Results; D013997:Time Factors