Three-photon excited fluorescence imaging of unstained tissue using a GRIN lens endoscope

We present a compact and portable three-photon gradient index (GRIN) lens endoscope system suitable for imaging of unstained tissues, potentially deep within the body, using a GRIN lens system of 1 mm diameter and 8 cm length. The lateral and axial resolution in water is 1.0 μm and 9.5 μm, respectively. The ~200 μm diameter field of view is imaged at 2 frames/s using a fiber-based excitation source at 1040 nm. Ex vivo imaging is demonstrated with unstained mouse lung at 5.9 mW average power. These results demonstrate the feasibility of three-photon GRIN lens endoscopy for optical biopsy.

[1]  W. Webb,et al.  Multifocal multiphoton endoscope. , 2012, Optics letters.

[2]  Watt W. Webb,et al.  In vivo imaging of unstained tissues using long gradient index lens multiphoton endoscopic systems , 2012, Biomedical optics express.

[3]  Eric J Seibel,et al.  Unique features of optical scanning, single fiber endoscopy * ** , 2002, Lasers in surgery and medicine.

[4]  F. Wise,et al.  Multiphoton microscopy system with a compact fiber‐based femtosecond‐pulse laser and handheld probe , 2011, Journal of biophotonics.

[5]  John White,et al.  Long-term two-photon fluorescence imaging of mammalian embryos without compromising viability , 1999, Nature Biotechnology.

[6]  Joseph C Liao,et al.  Optical biopsy of human bladder neoplasia with in vivo confocal laser endomicroscopy. , 2009, The Journal of urology.

[7]  Ina Pavlova,et al.  In vivo imaging of unstained tissues using a compact and flexible multiphoton microendoscope. , 2012, Journal of biomedical optics.

[8]  W. Webb,et al.  Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[9]  Ina Pavlova,et al.  Compact and flexible raster scanning multiphoton endoscope capable of imaging unstained tissue , 2011, Proceedings of the National Academy of Sciences.

[10]  W. Denk,et al.  Two-photon laser scanning fluorescence microscopy. , 1990, Science.

[11]  Shi-Wei Chu,et al.  Wavelength dependent damage in biological multi-photon confocal microscopy: A micro-spectroscopic comparison between femtosecond Ti:sapphire and Cr:forsterite laser sources , 2002 .

[12]  Xingde Li,et al.  Fiber-optic scanning two-photon fluorescence endoscope. , 2006, Optics letters.

[13]  W. Webb,et al.  Dual modality microendoscope with optical zoom capability , 2012, 2012 Conference on Lasers and Electro-Optics (CLEO).

[14]  N. Nishimura,et al.  Deep tissue multiphoton microscopy using longer wavelength excitation. , 2009, Optics express.

[15]  Riyi Shi,et al.  Characterization of photodamage in coherent anti-Stokes Raman scattering microscopy. , 2006, Optics express.

[16]  W. Denk,et al.  Two-photon imaging to a depth of 1000 microm in living brains by use of a Ti:Al2O3 regenerative amplifier. , 2003, Optics letters.

[17]  J. Fujimoto,et al.  Optical coherence tomography: an emerging technology for biomedical imaging and optical biopsy. , 2000, Neoplasia.

[18]  D. Kobat,et al.  In vivo two-photon microscopy to 1.6-mm depth in mouse cortex. , 2011, Journal of biomedical optics.

[19]  F. Helmchen,et al.  Ultra-compact fiber-optic two-photon microscope for functional fluorescence imaging in vivo. , 2008, Optics express.

[20]  Frank W. Wise,et al.  In vivo three-photon microscopy of subcortical structures within an intact mouse brain , 2012, CLEO 2012.

[21]  W. Webb,et al.  Measuring Serotonin Distribution in Live Cells with Three-Photon Excitation , 1997, Science.

[22]  K Bahlmann,et al.  Three-photon excitation in fluorescence microscopy. , 1996, Journal of biomedical optics.

[23]  V. Centonze,et al.  Three‐photon excitation fluorescence imaging of biological specimens using an all‐solid‐state laser , 1996 .

[24]  Woonggyu Jung,et al.  Design and implementation of fiber-based multiphoton endoscopy with microelectromechanical systems scanning. , 2009, Journal of biomedical optics.

[25]  Watt W. Webb,et al.  Use of a lensed fiber for a large-field-of-view, high-resolution, fiber-scanning microendoscope. , 2012, Optics letters.

[26]  Yuxin Leng,et al.  Scanning all-fiber-optic endomicroscopy system for 3D nonlinear optical imaging of biological tissues. , 2009, Optics express.

[27]  L. Fu,et al.  Three-dimensional nonlinear optical endoscopy. , 2007, Journal of biomedical optics.

[28]  W. Webb,et al.  Live tissue intrinsic emission microscopy using multiphoton-excited native fluorescence and second harmonic generation , 2003, Proceedings of the National Academy of Sciences of the United States of America.