We present a novel and inexpensive Stokes imaging spectropolarimeter based on the Snapshot Hyperspectral Imaging Fourier Transform (SHIFT) spectrometer. A rotating quarter wave plate and stationary linear polarizer placed in front of the SHIFT spectrometer enables us to reconstruct an object's spectra and Stokes parameters in the visible spectrum. Measurements are stored in the form of four-dimensional (4D) Stokes datacubes containing the object's spatial, spectral, and polarization information. We discuss calibration methods, review design considerations, and present preliminary results from proof-of-concept experiments.

We will present the concept, construction, and operation of Array Microscopes in the field of biomedical imaging. The benefits, technical challenges, and results of Array Microscope instruments will be discussed in detail.

An estimated 1.6 million breast biopsies are performed in the US each year. In order to provide real-time, in-vivo imaging with sub-cellular resolution for optical biopsies, we have designed an ultra-slim objective to fit inside the 1-mm-diameter hypodermic needles currently used for breast biopsies to image tissue stained by the fluorescent probe proflavine. To ensure high-quality imaging performance, experimental tests were performed to characterize fiber bundle's light-coupling efficiency and simulations were performed to evaluate the impact of candidate lens materials' autofluorescence. A prototype of NA = 0.4, 250-microm field of view, ultra-slim objective optics was built and tested, yielding diffraction-limited performance and estimated resolution of 0.9 microm. When used in conjunction with a commercial coherent fiber bundle to relay the image formed by the objective, the measured resolution was 2.5 microm.

This paper describes the design and fabrication of a novel array microscope for the first ultrarapid virtual slide processor (DMetrix DX-40 digital slide scanner). The array microscope optics consists of a stack of three 80-element 10 x 8-lenslet arrays, constituting a "lenslet array ensemble." The lenslet array ensemble is positioned over a glass slide. Uniquely shaped lenses in each of the lenslet arrays, arranged perpendicular to the glass slide constitute a single "miniaturized microscope." A high-pixel-density image sensor is attached to the top of the lenslet array ensemble. In operation, the lenslet array ensemble is transported by a motorized mechanism relative to the long axis of a glass slide. Each of the 80 miniaturized microscopes has a lateral field of view of 250 microns. The microscopes of each row of the array are offset from the microscopes in other rows. Scanning a glass slide with the array microscope produces seamless two-dimensional image data of the entire slide, that is, a virtual slide. The optical system has a numerical aperture of N.A.= 0.65, scans slides at a rate of 3 mm per second, and accrues up to 3,000 images per second from each of the 80 miniaturized microscopes. In the ultrarapid virtual slide processing cycle, the time for image acquisition takes 58 seconds for a 2.25 cm2 tissue section. An automatic slide loader enables the scanner to process up to 40 slides per hour without operator intervention. Slide scanning and image processing are done concurrently so that post-scan processing is eliminated. A virtual slide can be viewed over the Internet immediately after the scanning is complete. A validation study compared the diagnostic accuracy of pathologist case readers using array microscopy (with images viewed as virtual slides) and conventional light microscopy. Four senior pathologists diagnosed 30 breast surgical pathology cases each using both imaging modes, but on separate occasions. Of 120 case reads by array microscopy, there were 3 incorrect diagnoses, all of which were made on difficult cases with equivocal diagnoses by light microscopy. There was a strong correlation between array microscopy vs. "truth" diagnoses based on surgical pathology reports. The kappa statistic for the array microscopy vs. truth was 0.96, which is highly significant (z=10.33, p <0.001). There was no statistically significant difference between rates of agreement with truth between array microscopy and light microscopy (z=0.134, p >0.05). Array microscopy and light microscopy did not differ significantly with respect to the number/percent of correct decisions rendered (t=0.552, p=0.6376) or equivocal decisions rendered (t=2.449, p=0.0917). Pathologists rated 95.8% of array microscopy virtual slide images as good or excellent. None were rated as poor. The mean viewing time for a DMetrix virtual slide was 1.16 minutes. The DMetrix virtual slide processor has been found to reduce the virtual slide processing cycle more than 10 fold, as compared with other virtual slide systems reported to date. The virtual slide images are of high quality and suitable for diagnostic pathology, second opinions, expert opinions, clinical trials, education, and research.

The trade-off between field of view and resolution in conventional microscopy can be avoided by using multiple microscopes in parallel in the form of an array microscope. This paper explores the design, fabrication, and construction of such an array microscope. Imaging results using an array microscope will be presented.

Confocal microscopy provides images of tissue architecture and cellular morphology in living tissue in real time. Pilot clinical trials show confocal images can diagnose dysplasia with unprecedented specificity and sensitivity. Flexible fiber optic instrumentation permits in vivo imaging. This talk will overview confocal imaging for detection and management of cancer.

We present a fiber optic confocal reflectance microscope (FCRM) which can be used to image epithelial tissue with sub-cellular resolution in vivo. Confocal images of normal and abnormal appearing cervical tissue were obtained in vivo from eighteen patients undergoing colposcopic examination of the cervix; biopsy specimens were taken from imaged sites. The measured lateral and axial resolutions of the system were 1.6 microm and 3 microm, respectively. Morphologic features, including nuclear size and nuclear-to-cytoplasmic ratio, were extracted from confocal images obtained at various depths beneath the epithelial surface. Image features extracted from confocal images compared well with features extracted from confocal images obtained in vitro and from previous histopathologic studies. This study shows that fiber optic confocal reflectance microscopy can be used to visualize the morphology of cervical epithelium in vivo.

In collaboration with the Department of Biomedical Engineering at the University of Texas at Austin and the UT MD Anderson Cancer Center, a laser scanning fiber confocal reflectance microscope (FCRM) system has been designed and tested for in vivo detection of cervical and oral pre-cancers. This system along with specially developed diagnosis algorithms and techniques can achieve an unprecedented specificity and sensitivity for the diagnosis of pre-cancers in epithelial tissue.(1) The FCRM imaging system consists of an NdYAG laser (1064 run), scanning mirrors/optics, precision pinhole, detector, and an endoscopic probe (the objective). The objective is connected to the rest of the imaging system via a fiber bundle. The fiber bundle allows the rest of the system to be remotely positioned in a convenient location. Only the objective comes into contact with the patient. It is our intent that inexpensive mass-produced disposable endoscopic probes would be produced for large clinical trails. This paper touches on the general design process of developing a miniature, high numerical aperture, injection-molded (IM) objective. These IM optical designs are evaluated and modified based on manufacturing and application constraints., Based on these driving criteria, one specific optical design was chosen and a detailed tolerance analysis was conducted. The tolerance analysis was custom built to create a realistic statistical analysis for integrated IM lens elements that can be stacked one on top of another using micro-spheres resting in tiny circular grooves. These configurations allow each lens element to be rotated and possibly help compensate for predicted manufacturing errors. This research was supported by a grant from the National Institutes of Health (ROI CA82880). Special thanks go to Applied Image Group/Optics for the numerous fabrication meetings concerning the miniature IM objective.

An endoscopic confocal microscope requires a high-performance, miniaturized microscope objective. We present the design of a miniature water-immersion microscope objective that is approximately 10 times smaller in length than a typical commercial objective. The miniature objective is 7 mm in outer diameter and 21 mm in length (from object to image). It is used in a fiber confocal reflectance microscope. The miniature microscope objective has a numerical aperture of 1.0, a field of view of 250 microm, and a working distance of 450 microm. It delivers diffraction-limited performance at lambda = 1064 nm. Micrometer-level resolution has been experimentally demonstrated.

We have built a fiber optic confocal reflectance microscope capable of imaging human tissue at 15 frames/s. Miniaturization of the objective lens and the mechanical component for positioning and axially scanning the objective enable the system to be used in inner organs of the human body. The measured lateral resolution is 2 micrometers and axial resolution is 10 micrometers. Confocal images of human lip in vivo have been obtained without any fluorescent dyes. Cell nuclear morphology and tissue architecture can be appreciated from these images. This microscope has the potential for in situ detection of pathology.

In this paper we present the design and construction of a NA=1.0 water- immersion microscope objective 5mm in diameter and 17mm in length. Proposed design can be fabricated by injection molding using plastic materials.

In this paper, we describe the progress toward the development of miniaturized imaging systems with applications in medical imaging, and specifically, detection of pre-cancer. The focus of the article is a miniature, optical-sectioning, fluorescence microscope. The miniature microscope is constructed from lithographically printed optics and assembled using a bulk micro-machined silicon microoptical table. Optical elements have been printed in a negative tone hybrid glass to a maximum depth of 59 /spl mu/m and an rms surface roughness between 10-45 nm, fulfilling the requirements of the miniature microscope. Test optical elements have been assembled using silicon-spring equipped mounting slots. The design of silicon springs is presented in this paper. Optical elements can be assembled within the tolerances of an NA=0.4 miniature microscope objective, confirming the concept of simple, zero-alignment assembly.