We present the design and feasibility testing of a multimodal co-registered endoscope based on a dual-path optical system integrated with a scanning piezo. This endoscope incorporates three different imaging modalities. A large field-of-view (FOV) reflectance imaging system enables visualization of objects several millimeters in front of the endoscope, while optical coherence microscopy (OCM) and multiphoton microscopy (MPM) are employed in contact with tissue to further analyze suspicious areas. The optical system allows multiple different imaging modalities by employing a dual optical path. One path features a low numerical aperture (NA) and wide FOV to allow reflectance imaging of distant objects. The other path features a high NA and short working distance to allow microscopy techniques such as OCM and MPM. Images of test targets were obtained with each imaging modality to verify and characterize the imaging capabilities of the endoscope. The reflectance modality was demonstrated with a 561 nm laser to allow high contrast with blood vessels. It achieved a lateral resolution of 24.8 μm at 5 mm and a working distance from 5 to 30 mm. OCM was performed with a 1300 nm super-luminescent diode since this wavelength experiences low relative scattering to allow for deeper tissue imaging. Measured OCM lateral and axial resolution was 4.0 and 14.2 μm, respectively. MPM was performed with a custom 1400 nm femtosecond fiber laser, a wavelength suitable for exciting multiple exogenous, and some endogenous fluorophores, as well as providing information on tissue composition through harmonic generation processes. A 4.0 μm MPM lateral resolution was measured.
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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