Early indicators of cancer manifest as cellular abnormalities that can be imaged via high resolution imaging modalities such as label-free Multiphoton Microscopy (MPM). Implementing MPM endoscopically can aid in the early diagnosis of cancer. We are developing a minimally invasive microendoscope system capable of simultaneous co-registered multiphoton imaging (two- and three-photon excited fluorescence, second and third harmonic generation) of the epithelial layer in small diameter ductal tissues via helical scanning of a 1.0mm diameter endoscope distal end with a fixed focus. The endoscope working length is comprised of a stationary outer sheath housing a proximally driven endoscope distal optical system. The lenses are 0.5mm in diameter and side viewing, requiring a novel optical design with power on the exit surface of the fold prism. Additive manufacturing (three-dimensional [3D] printing) opens significantly more possibilities for distal end microendoscope optical design. We present the design of the distal end outer sheath, housing, and optics, as well as an evaluation of the feasibility of 3D printed optics for a high numerical aperture (HNA) MPM microendoscope system. The selected distal end outer sheath meets flexibility, size, and optical requirements suitable for a first iteration lab prototype. The distal end housing was custom designed to be easily attached and removed from the proximal system and to ensure stable and consistent helical motion of the inner probe distal end when it is flexed and bent in manners needed for use in curved tissue lumens. We have also designed and manufactured (via two photon polymerization) multiple configurations of a 0.5mm monolithic multi-element lens system containing an aspheric surface on the exit face of the prism and evaluated its optical performance.
Early detection of cancer is crucial for improving patient survival. High resolution optical imaging is ideal to image cellular abnormalities indicative of early cancer. For tissues located deep within the body, such as the pancreato-biliary ducts, high resolution imaging must be implemented endoscopically due to the limited penetration depth of light. We are developing a minimally invasive high numerical aperture (HNA) microendoscope system capable of simultaneous co-registered multiphoton imaging (two-photon excited fluorescence, second harmonic generation, three-photon excited fluorescence, and third harmonic generation) of small diameter ductal tissues, such as the pancreato-biliary ducts. Imaging of the epithelial layer is achieved via helical scanning of the 1.5 mm diameter endoscope with a fixed focus. The endoscope distal end optics act as both the illumination and collection mechanism, with the core of the dual clad fiber (DCF) carrying femtosecond laser excitation light, and the inner cladding of the DCF carrying multiphoton emission. Designing HNA optics at the 1 mm diameter size scale is challenging, time consuming, and may be expensive. To complete development of the proximal components of the system, we designed a low numerical aperture (LNA) reflectance & single photon fluorescence system using low cost off the shelf optical components to aid in the development of software and the testing of proximal system hardware components. Additionally, rapid, low-cost design and fabrication of HNA optics with 3D printing is presented.
Multimodal imaging is an advantageous method to increase the accuracy of disease classification. As an example, we and others have shown that optical coherence tomography images and fluorescence spectroscopy contain complementary information that can increase the sensitivity and specificity for cancer detection. A common challenge in multimodal imaging is image co-registration. The different images are often taken with separate imaging setups, making it challenging to precisely image the same tissue area or co-register the images computationally. To solve this problem, we have developed a co-registered multimodal imaging system that images the same tissue location with reflectance, multi-photon, and optical coherence microscopy. The co-registration mechanism is a dual-clad fiber that integrates with a scanning microscope or scanning endoscope, collecting all three signals using the same optical path. In the current implementation, optical coherence tomography utilizes a 1300 nm super luminescent diode, multi-photon signals are excited by a custom femtosecond 1400 nm fiber laser producing two- and three-photon signals in the 460-900 nm band, and reflectance imaging operates at 561 nm. The system separates the different signals using fiber wavelength division multiplexers, a dual-clad fiber coupler, and dichroic mirrors to deliver the different signals to the corresponding detector. This wavelength selection enables the system to work passively, meaning that there is no need for devices such as filter wheels. Using the scanning microscope configuration, we have obtained multimodal images of ex-vivo ovine ovary tissue.
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