Development of a real-time flexible multiphoton microendoscope for label-free imaging in a live animal

G. Ducourthial , P. Leclerc , T. Mansuryan , M. Fabert , J. Brevier , R. Habert , F. Braud , R. Batrin , C. Vever-Bizet , G. Bourg-Heckly , L. Thiberville , A. Druilhe , A. Kudlinski , F. Louradour

Bibtex , URL
Sci Rep, 5, 18303
Published 17 Dec. 2015
DOI: 10.1038/srep18303
ISSN: 2045-2322


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 mu m and 12 mu m, with a field-of-view as large as 450 mu m. 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 mu m 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.

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