Andrew R. Rouse

Slit-scanning confocal microendoscope for high-resolution in vivo imaging

We discuss the design and construction of a novel imaging system in which a fiber-optic imaging bundle and miniature optical and mechanical components are used to allow confocal fluorescence microscopy in remote locations. The instrumentation has been developed specifically for cellular examination of tissue for optical biopsy. Miniaturization of various components makes the device usable in a clinical setting. The numerical aperture of the beam in the tissue is 0.5, and the field of view is 430 microm. The measured lateral resolution of the system is 3.0 microm. The axial point and the axial planar response functions of the confocal system were measured with a FWHM of 10 and 25 microm, respectively. In vitro and in vivo images obtained with cell cultures, human tissue specimens, and animal models indicate that the performance of the device is adequate for microscopic evaluation of cells.

Design and demonstration of a miniature catheter for a confocal microendoscope

The fluorescence confocal microendoscope provides high-resolution, in vivo imaging of cellular pathology during optical biopsy. The confocal microendoscope employs a flexible fiber-optic catheter coupled to a custom-built slit-scan confocal microscope. The catheter consists of a fiber-optic imaging bundle linked to a miniature objective and focus assembly. The 3-mm-diameter catheter may be used on its own or routed though the instrument channel of a commercial endoscope, adding microscopic imaging capability to conventional endoscopy. The design and performance of the miniature objective and focus assembly are discussed. Primary applications of the system include diagnosis of disease in the gastrointestinal tract and female reproductive system.

Multispectral imaging with a confocal microendoscope.

The concept of a multispectral confocal microscope for in vivo imaging is introduced. To demonstrate the concept we modified a slit-scan fluorescence confocal microendoscope incorporating a fiber-optic catheter for in vivo imaging to record multispectral images. The system was designed to examine cellular structures during optical biopsy and to exploit the diagnostic information contained within the spectral domain. Preliminary experiments were carried out in phantoms and cell cultures to demonstrate the potential of the technique.

Clinical confocal microlaparoscope for real-time in vivo optical biopsies

Successful treatment of cancer is highly dependent on the stage at which it is diagnosed. Early diagnosis, when the disease is still localized at its origin, results in very high cure rates-even for cancers that typically have poor prognosis. Biopsies are often used for diagnosis of disease. However, because biopsies are destructive, only a limited number can be taken. This leads to reduced sensitivity for detection due to sampling error. A real-time fluorescence confocal microlaparoscope has been developed that provides instant in vivo cellular images, comparable to those provided by histology, through a nondestructive procedure. The device includes an integrated contrast agent delivery mechanism and a computerized depth scan system. The instrument uses a fiber bundle to relay the image plane of a slit-scan confocal microlaparoscope into tissue. It has a 3-mum lateral resolution and a 25-mum axial resolution. Initial in vivo clinical testing using the device to image human ovaries has been done in 21 patients. Results indicate that the device can successfully image organs in vivo without complications. Results with excised tissue demonstrate that the instrument can resolve sufficient cellular detail to visualize the cellular changes associated with the onset of cancer.

Multispectral confocal microendoscope for in vivo and in situ imaging

We describe the design and operation of a multispectral confocal microendoscope. This fiber-based fluorescence imaging system consists of a slit-scan confocal microscope coupled to an imaging catheter that is designed to be minimally invasive and allow for cellular level imaging in vivo. The system can operate in two imaging modes. The grayscale mode of operation provides high resolution real-time in vivo images showing the intensity of fluorescent signal from the specimen. The multispectral mode of operation uses a prism as a dispersive element to collect a full multispectral image of the fluorescence emission. The instrument can switch back and forth nearly instantaneously between the two imaging modes (less than half a second). In the current configuration, the multispectral confocal microendoscope achieves 3-microm lateral resolution and 30-microm axial resolution. The system records light from 500 to 750 nm, and the minimum resolvable wavelength difference varies from 2.9 to 8.3 nm over this spectral range. Grayscale and multispectral imaging results from ex-vivo human tissues and small animal tissues are presented.

Computer-aided identification of ovarian cancer in confocal microendoscope images.

The confocal microendoscope is an instrument for imaging the surface of the human ovary. Images taken with this instrument from normal and diseased tissue show significant differences in cellular distribution. A real-time computer-aided system to facilitate the identification of ovarian cancer is introduced. The cellular-level structure present in ex vivo confocal microendoscope images is modeled as texture. Features are extracted based on first-order statistics, spatial gray-level-dependence matrices, and spatial-frequency content. Selection of the features is performed using stepwise discriminant analysis, forward sequential search, a nonparametric method, principal component analysis, and a heuristic technique that combines the results of these other methods. The selected features are used for classification, and the performance of various machine classifiers is compared by analyzing areas under their receiver operating characteristic curves. The machine classifiers studied included linear discriminant analysis, quadratic discriminant analysis, and the k-nearest-neighbor algorithm. The results suggest it is possible to automatically identify pathology based on texture features extracted from confocal microendoscope images and that the machine performance is superior to that of a human observer.

Monte Carlo characterization of parallelized fluorescence confocal systems imaging in turbid media

We characterize and compare the axial and lateral performance of fluorescence confocal systems imaging in turbid media. The aperture configurations studied are a single pinhole, a slit, a Nipkow disk, and a linear array of pinholes. Systems with parallelized apertures are used clinically because they enable high-speed and real-time imaging. Understanding how they perform in highly scattering tissue is important. A Monte Carlo model was developed to characterize parallelized system performance in a scattering media representative of human tissues. The results indicate that a slit aperture has degraded performance, both laterally and axially. In contrast, the analysis reveals that multipinhole apertures such as a Nipkow disk or a linear pinhole array can achieve performance nearly equivalent to a single pinhole aperture. The optimal aperture spacing for the multipinhole apertures was determined for a specific tissue model. In addition to comparing aperture configurations, the effects of tissue nonradiative absorption, scattering anisotropy, and fluorophore concentration on lateral and axial performance of confocal systems were studied.

Dual modality fluorescence confocal and spectral-domain optical coherence tomography microendoscope

Optical biopsy facilitates in vivo disease diagnoses by providing a real-time in situ view of tissue in a clinical setting. Fluorescence confocal microendoscopy and optical coherence tomography (OCT) are two methods that have demonstrated significant potential in this context. These techniques provide complementary viewpoints. The high resolution and contrast associated with confocal systems allow en face visualization of sub-cellular details and cellular organization within a thin layer of biological tissue. OCT provides cross-sectional images showing the tissue micro-architecture to a depth beyond the reach of confocal systems. We present a novel design for a bench-top imaging system that incorporates both confocal and OCT modalities in the same optical train allowing the potential for rapid switching between the two imaging techniques. Preliminary results using simple phantoms show that it is possible to realize both confocal microendoscopy and OCT through a fiber bundle based imaging system.

Development of a fiber-optic confocal microendoscope for clinical endoscopy

A confocal micro endoscope has been developed to examine cellular pathology during optical biopsy. The system employs a flexible fiber optic catheter coupled to a slit-scan confocal microscope to image tissue at remote locations in the body. The catheter of the confocal micro endoscope consists of a fiber-optic imaging bundle, a miniature objective, and a miniature focusing mechanism. The system has a lateral resolution of 1.8 micrometers and an axial resolution of 25 micrometers . The confocal micro endoscope can operate in a grayscale or multi-spectral imaging mode. Extensive work has been done to design a new miniature objective and focusing mechanism that will allow the catheter to be routed through the therapeutic channel of a clinical endoscope. We present the design for a miniature F/1 achromatic objective with nearly diffraction limited performance. The objective will be coupled to a pneumatic focusing mechanism to provide focus control to 200 micrometers below the surface of the tissue. The new catheter has an overall diameter of 3mm with a rigid tip of only 20mm in length.

Automated texture-based identification of ovarian cancer in confocal microendoscope images

The fluorescence confocal microendoscope provides high-resolution, in-vivo imaging of cellular pathology during optical biopsy. There are indications that the examination of human ovaries with this instrument has diagnostic implications for the early detection of ovarian cancer. The purpose of this study was to develop a computer-aided system to facilitate the identification of ovarian cancer from digital images captured with the confocal microendoscope system. To achieve this goal, we modeled the cellular-level structure present in these images as texture and extracted features based on first-order statistics, spatial gray-level dependence matrices, and spatial-frequency content. Selection of the best features for classification was performed using traditional feature selection techniques including stepwise discriminant analysis, forward sequential search, a non-parametric method, principal component analysis, and a heuristic technique that combines the results of these methods. The best set of features selected was used for classification, and performance of various machine classifiers was compared by analyzing the areas under their receiver operating characteristic curves. The results show that it is possible to automatically identify patients with ovarian cancer based on texture features extracted from confocal microendoscope images and that the machine performance is superior to that of the human observer.