Research On Fiber Bundle Based Coherent Anti-stokes Raman Scattering Endomicroscopy System

Coherent anti-Stokes Raman scattering（CARS）imaging is a label-free chemicallyselective imaging modality, and is suitable for applications in biomedical imaging. CARSpossesses many merits due to its nonlinear and coherent nature, such as label-free chemicallyselective imaging based on molecular vibration, fast imaging speed, high sensitivity, threedimensional sectioning imaging and submicron resolution. CARS microscopy is already amatured technology, while its clinical application on in vivo imaging requires thedevelopment of CARS endomicroscope. Miniaturization is currently the most challengingproblem in its development. To miniaturize the system and achieve high collection efficiencyof CARS photons, we applied fiber bundle into CARS endomicroscopy system. There arefour parts in this research:1. A fiber bundle based CARS endomicroscopy system was built, and four-wave-mixing(FWM) noise suppression based on polarization control and dual-wavelength waveplate wereused to acquire pure CARS image without FWM noise. The CARS photon collectionefficiency of fiber bundle was measured, and it is much higher than that of single excitationfiber. CARS images of mouse adipocytes and lung tissue were acquired with the fiber bundlecollection. The3D sectioning imaging ability and CARS spectroscopy ability of the systemwas also proved by mouse adipocyte imaging. The availability of fiber bundle collection inCARS endomicroscopy system was demonstrated.2. A dual channel fiber bundle based endomicroscopy system for simultaneous CARSand SHG imaging was designed and demonstrated. SHG imaging ability was added into fiberbundle based CARS endomicroscopy system. The CARS photon collection efficiency of thetwo channels were measured. The difference between the images collected by the twochannels was systematically studied and explained. CARS images of mouse adipocyte wereacquired with the two channels, and SHG images of mouse tail tendon were also acquired.The simultaneous CARS and SHG imaging ability of the system was demonstrated byimaging a mouse tail tissue.3. The CARS photon collection performances of the single excitation fiber and the fiberbundle were systematically compared. A method based on simultaneous single fiber and fiberbundle collection to differentiate epi-photon and forward-photon images was proposed. AZEMAX model of the system was built to compare the distribution of fiber collection lightcones at the focal plane of the60X objective, and the collection of epi-and forward-CARS photons by the single fiber and the fiber bundle was discussed. CARS images of mouseadipocytes collected by the single fiber and the fiber bundle were compared. For SHGimaging, a mathematical model was built to describe the collection of epi-and forward-SHGphotons by the single fiber and the fiber bundle. Epi-and forward-SHG images of mouse tailtendon were separated with this model, and the ratio of these two types of photons collectedby the single fiber was calculated. A structural distribution pattern of epi-and forward-SHGphoton scatterers in mouse tail tendon was observed with this method, and that pattern isrelated to the structure of mouse tail tendon. With this system, the images of epi-and forward-CARS or SHG photons of thick tissues were differentiated for the first time, and the imagesof these two types of photons are very important for biological studies.4. The piezo tube scanning method was combined with the fiber bundle collection scheme.A CAR/SHG endomicroscopy system based on piezo tube scanning and fiber bundlecollection was built. The fiber bundle was driven by the piezo tube for scanning, the size ofthe scanning part of fiber bundle based CARS endomicroscopy system was miniaturized, andthe scanning speed was improved. CARS images of mouse adipocytes and SHG images ofmouse tail tendon were obtained with this system. The ability of the piezo tube to drive thefiber bundle and the fast imaging ability of the system were demonstrated.This research is a foundation for the application of fiber bundle in CARS and SHGendomicroscopy system, and is also an exploration for using fiber bundle to achieve theminiaturization of CARS endomicroscopy system. This research also contributs to thebiomedical application of the fiber bundle based CARS endomicroscopy system.