Considerable improvement in image quality and ease of use of the original clinical instrument was accomplished with the design of an improved objective lens by Laing that resulted in images of sufficient quality, even in edematous corneas, that were suitable for morphometric analysis using computerized image processing systems developed by Laing and associates10. Various methods have been used to obtain wider fields of cells than are obtained using a stationary slit of light. Sherrard and associates11 developed a fluorite cone for the objective lens that reduced the interfering objective lens-epithelial reflection and enabled a wider slit to be used that resulted in a wider field of cells imaged, at least in clear corneas that have minimal light scatter from the stroma. Koester12 designed an optical system that moved a narrow slit of light repeatedly up an down on the endothelium in such a way that a wide image was produced without suffering degradation from stromal scattering. Lemp and associates13 demonstrated that a Petran confocal microscope14,15 was capable of obtaining endothelial, epithelial, and stromal images not previously possible. With the general increase in interest in confocal microscope techniques (resulting, in part, from the expiration of the Petran patent on the rotating disk confocal microscope), a variety of rotating disk and rotating slit specular microscopes have been developed.12,16,17 Such complex instruments have demonstrated considerably improved images of nerve fibers, keratocytes and other stromal structures, although images of the endothelium are not appreciably improved over that which is obtained using simpler and more reliable and less expensive stationary slit specular microscopes.

The Bio-Optics LSM-12000 is the most automatic instrument yet developed; tracking of the cornea and imaging the endothelium is fully automatic, requiring minimal intervention by the operator. In using this instrument, the patient is instructed to place their chin onto either the left or the right chin rest, depending upon whether photographs of the left or the right eye are desired, and to look at the green fixation light. The operator needs only to adjust the chin rest vertically and perhaps move the head slightly to the right or the left until the pupil of the eye, seen on the video monitor on the front of the instrument, is approximately centered. The operator then presses a red button on the control box to start the automated photography process. The optics of the instrument first objectively aligns itself relative to the cornea by using the Purkinke images until the proper specular reflection mode is achieved. The instrument then objectively focusses back to the endothelial surface, the green fixation light begins to flash on and off (indicating that the patient should not blink), the flash lamp is triggered, and the relulting endothelial photograph is displayed on the video monitor screen. Unlike the other clinical specular microscopes that are available, the LSM-12000 requires essentially no training of the operator. Patients can even take their own endothelial photographs by looking into the instrument and pressing the red button to start the automated photography process. The instrument has built into it the capability of doing a fixed frame cell count. An advanced model also has the ability of photographing the peripheral endothelium, 3 mm from the center of the cornea at the 12, 2, 6, and 10 o'clock positions, as well as the capability of doing individual cell analysis using a nearest-neighbor method that gives approximate values for the percent of hexagonal cells, average area, and coefficient of variation of area in a contiguous group of cells whose centers are digitized using the mouse that comes with the instrument. The instrument has a video output that can input the image into an image storage device or into an advanced endothelial cell analysis system such as the Bambi system (Bio-Optics) for more standard cell counting and analysis, image and data management, and for digital image and data storage.

References:

17. Kino G, Chore C, Xiao Q. Imaging theory for the scanning optical microscope. Proc-SPIE. 1988;1028:104.

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