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The purpose of this study was to evaluate the retina using near-infrared (NIR) two-photon scanning laser ophthalmoscopy. New Zealand white rabbits, albino rats, and brown Norway rats were used in this study. An autofluorescence image of the retina, including the retinal cells and its associated vasculatures was obtained by a real-time scan using the ophthalmoscope. Furthermore, the retinal vessels, nerve fiber layers and the non-pigmented retina were recorded with two-photon fluorescein angiography (FA); and the choroidal vasculatures were recorded using two-photon indocyanine green angiography (ICGA). Two-photon ICGA was achieved by exciting a second singlet state at ∼398 nm. Simultaneous two-photon FA and two-photon ICGA were performed to characterize the retinal and choroidal vessels with a single injection. The minimum laser power threshold required to elicit two-photon fluorescence was determined. The two-photon ophthalmoscope could serve as a promising tool to detect and monitor the disease progression in animal models. Moreover, these high-resolution images of retinal and choroidal vessels can be acquired in a real-time scan with a single light source, requiring no additional filters for FA or ICGA. The combination of FA and ICGA using the two-photon ophthalmoscope will help researchers to characterize the retinal diseases in animal models, and also to classify the types (classic, occult or mixed) of choroidal neovascularization (CNV) in macular degeneration. Furthermore, the prototype can be adapted to image the retina of rodents and rabbits.

Adaptive optics has emerged as an empowering technology for retinal imaging with cellular resolution. This technology holds potential for non-invasive detection and diagnoses of the most common eye diseases such as glaucoma, diabetic retinopathy and age-related macular degeneration (AMD). However, the cost and complexity of adaptive optics ophthalmoscopes currently impede its clinical use. A scanning laser ophthalmoscope with phase plate has the potential to establish adaptive optics in a clinical application by simplifying its incorporation in the prevailing systems. Phase plates can correct aberrations down to diffraction limit and can be easily incorporated into ophthalmoscopes to improve the retinal image quality.

More than 25 years ago, the first in vivo measurement of the eye’s wave aberration was demonstrated at the University of Heidelberg, using the Shack-Hartmann method. The continuous measurement of ocular aberrations paved the way to employ adaptive optics for the correction of ocular aberrations in ophthalmoscopy. Various new diagnostic and treatment modalities have since been successfully introduced, e.g. adaptive optics scanning laser ophthalmoscopy (AO-SLO), adaptive optics optical coherence tomography (AO-OCT) and adaptive optics two-photon ophthalmoscopy (AO-TPO). Employing adaptive optics techniques, the limits of human vision can be assessed. With the development of wavefront-guided laser refractive surgery, the quest for optically perfect vision is being pursued over the last 20 years.

Cataract surgeons are in constant pursuit to achieve optimal visual outcomes for their patients. Therefore, numerous technologies and techniques have emerged, most of which are designed to assist the surgeon preoperatively and intraoperatively. The in vivo modification is achieved by creating a lens within an existing IOL using a femtosecond laser. The lens within the lens is created by altering the hydrophilicity of an existing and standard IOL in certain designated areas. The changes created by the perfect lens technology alter the IOL’s refractive outcome and enable the surgeon to create the patient’s desired vision in a simple postoperative in-office procedure.

Femtosecond-Laser-Assisted cataract surgery (FLACS) offers surgeons a reproducible, non-invasive technique to replace the least predictable and technically most demanding steps of conventional cataract procedures. A computer-guided laser linked to an optical imaging system (e.g. OCT) performs the corneal incision, capsulotomy, and lens fragmentation steps, thus changing the requirements associated with traditional techniques by removing the need for blade incisions and reducing phacoemulsification time and energy. Initial research has confirmed appropriate safety and efficacy of Femtosecond-Laser-Assisted cataract surgery, with improvements in anterior capsulotomy, phacofragmentation, and corneal incision. The potential gains in precision associated with FLACS may improve safety and clinical outcomes.

Since the invention of the first ophthalmoscope in 1851, the quest for quantitative in vivo imaging of ocular surfaces for diagnostic purposes was initiated. Confocal microscopy has been widely implemented in clinics providing excellent spatial resolution and enabling optical sectioning. With the introduction of ultrashort pulsed laser sources, multiphoton microscopy became feasible. Multiphoton microscopy allows 3D tissue imaging. Collagen, elastin, NAD(P)H etc., can be imaged by multiphoton microscopy, providing structural and functional information of the cornea. In this article, we will discuss the principles and potential use and limitations of confocal and multiphoton microscopy for corneal imaging.

More than 25 years ago, the first in vivo measurement of the eye’s wave aberration was demonstrated at the University of Heidelberg, using the Shack-Hartmann method. Since then, various new diagnostic and treatment modalities have been successfully introduced, like adaptive optics scanning laser ophthalmoscopy (AO-SLO), adaptive optics optical coherence tomography (AO-OCT) and adaptive optics two-photon ophthalmoscopy (AO-TPO), as well as customized refractive-surgical techniques, like customized LASIK (e.g. CustomVue), femtosecond laser refractive surgery (e.g. ReLEx, SMILE), femtosecond laser assisted cataract surgery (FLACS) and various customized intraocular lens (IOL) developments, including postoperative in-vivo IOL fine-tuning. Article not available.

Imaging of non-linear optical (NLO) signals generated from the eye using ultrafast pulsed lasers has been limited to the study of ex vivo tissues because of the use of conventional microscopes with slow scan speeds. The purpose of this study was to evaluate the ability of a novel, high scan rate ophthalmoscope to generate NLO signals using an attached femtosecond laser. NLO signals were generated and imaged in live, anesthetized albino rabbits using a newly designed Heidelberg Two-Photon Laser Ophthalmoscope with attached 25 mW femtosecond laser having a central wavelength of 780 nm, pulsewidth of 75 fs, and a repetition rate of 50 MHz. To assess two-photon excited fluorescent (TPEF) signal generation, cultured rabbit corneal fibroblasts (RCF) were first labeled by Blue-green fluorescent FluoSpheres (1 μm diameter) and then cells were micro-injected into the central cornea. Clumps of RCF cells could be detected by both reflectance and TPEF imaging at 6 hours after injection. By 6 days, RCF containing fluorescent microspheres confirmed by TPEF showed a more spread morphology and had migrated from the original injection site. Overall, this study demonstrates the potential of using NLO microscopy to sequentially detect TPEF signals from live, intact corneas. We conclude that further refinement of the Two-photon laser Ophthalmoscope should lead to the development of an important, new clinical instrument capable of detecting NLO signals from patient corneas.

A compact adaptive optics system was developed and integrated into the system for multiphoton laser scanning ophthalmoscopy. Defocus, astigmatism, trefoil, and coma are compensated via a combination of telescope and rotating pairs of phase plates. The spherical aberration as well as all dynamic aberrations are corrected by an active micro-electromechanical (MEMS) mirror. Wavefront measurement is conducted by fast SHS and pyramid sensor devices. The MEMS-mirror based adaptive optics system was validated in a study on 60 eyes of 30 subjects.The adaptive-optical multiphoton ophthalmoscope is aimed at detecting the early onset of retinal diseases such as e.g. age-related macula degeneration.

Purpose To assess the efficiency and effectiveness of the technology that creates a hydrophilicity-based refractive index change within a standard intraocular lens (IOL) to alter the refractive characteristics of the IOL. Setting Perfect Lens LLC, Irvine, California, USA. Design Experimental study. Methods The IOL used in this experiment was a standard hydrophobic model (EC-1Y). The refractive index of the material was changed by exposure of the material to a femtosecond laser and the subsequent absorption of water by the material. An experimental system using a femtosecond laser, an acoustic-optic modulator, beam-shaping optics, a scan system, and an objective lens was used to create the refractive index change within the IOL. Experiments were performed to determine the optimum wavelength, energy per pulse, and line spacing to produce the refractive index shaping lens. A power and modulation transfer function (MTF) measurement device for refractive and diffractive IOLs was used to measure the diopter and MTF before and after the creation of the refractive index shaping lens. Results The technology successfully altered the refractive characteristics of the IOL. The refractive index change altered the diopter (D) of the IOL (to within ±0.1 D of the targeted change) without significant diminution in the MTF ( Conclusion The refractive properties of an IOL can be altered by building a refractive index shaping lens within an IOL using a femtosecond laser with minimal diminution in MTF. Financial Disclosure All authors are employed by Perfect Lens, LLC and have a financial interest in the products of Perfect Lens, LLC.

The chemical basis for the alteration of the refractive properties of an intraocular lens with a femtosecond laser was investigated. Three different microscope setups have been used for the study: Laser Induced Fluorescence (LIF) microscopy, Raman microscopy and coherent anti-Stokes Raman Scattering (CARS) microscopy. Photo-induced hydrolysis of polymeric material in aqueous media produces two hydrophilic functional groups: acid group and alcohol group. The spectral signatures identify two of the hydrophilic polar molecules as N-phenyl-4-(phenylazo)-benzenamine (C18H15N3) and phenazine-1-carboxylic acid (C13H8N2O2). The change in hydrophilicity results in a negative refractive index change in the laser-treated areas.

Adaptive optics and femtosecond laser surgical systems have allowed the measurement and correction of the optical properties of the human eye, extending the knowledge of the limits of human vision.

We investigated the use of ultrashort pulsed (femtosecond) laser technology in corneal refractive surgery. When compared to longer pulsewidth nanosecond or picosecond laser pulses, femtosecond laser-tissue interactions are characterized by significantly smaller and more deterministic photodisruptive energy thresholds, as well as reduced shock waves and smaller cavitation bubbles. We utilized a highly reliable all-solid-state femtosecond laser system for all studies to demonstrate practicality in real-world operating conditions. Contiguous tissue effects were achieved by scanning a 5-/spl mu/m focused laser spot below the corneal surface at pulse energies of approximately 2-4 /spl mu/J. A variety of scanning patterns was used to perform three prototype procedures in animal eyes; corneal flap cutting, keratomileusis, and intrastromal vision correction. Superior dissection and surface quality results were obtained for lamellar procedures (corneal flap cutting and keratornileusis). Preliminary in vivo studies of intrastromal vision correction suggest that consistent refractive changes can also be achieved with this method. We conclude that femtosecond laser technology may be able to perform a variety of corneal refractive procedures with high precision, offering advantages over current mechanical and laser devices and techniques.

Laser scanning tomography can be used to assess retinal nerve fiber layer thickness and optic disc topography of the human eye. A pinhole is located at a plane conjugate to the focal plane of the scanning laser beam. This so-called confocal configuration assures that only the light originating from the illuminated focal plane on the retina passes through the pinhole and is detected by the photomultiplier. Consequently, images with high spatial resolution in all directions are obtained. An active optical system (active mirror) further improves the lateral/depth resolution of the laser tomographic scanner. By partially compensating for the optical aberrations introduced by the cornea and lens, the active optical system allows the illuminating beam to be enlarged to 6 mm, thus improving depth resolution twofold.

Collagen, as the most abundant protein in the human body, determines the unique physiological and optical properties of the connective tissues including cornea and sclera. The ultrastructure of collagen, which conventionally can only be resolved by electron microscopy, now can be probed by optical second harmonic generation (SHG) imaging. SHG imaging revealed that corneal collagen fibrils are regularly packed as a polycrystalline lattice, accounting for the transparency of cornea. In contrast, scleral fibrils possess inhomogeneous, tubelike structures with thin hard shells, maintaining the high stiffness and elasticity of the sclera.

Age-related macular degeneration (AMD) is among the major concerns in ophthalmology, as it is the primary cause for irreversible blindness in developed countries. Nevertheless, there is poor understanding of the origins and mechanisms that trigger this important ocular disease. In common clinical pratice, AMD is monitored by autofluorescence imaging of the retinal pigment epithelial (RPE) cells through a confocal scanning laser ophthalmoscope. The RPE cells derive their dominant autofluorescence from the lipofuscin granules that accumulate in the cytoplasm with increasing age and disease. We explored a different approach to retinal RPE imaging using two-photon excited autofluorescence, offering intrinsic three-dimensional resolution, larger sensing depth and reduced photodamage compared to single-photon excited fluorescence ophthalmoscopy. A two-photon microscope, based on the architecture of a conventional scanning laser ophthalmoscope (HRT, Heidelberg Engineering, Germany), was designed for autofluorescence imaging on retina samples from postmortem human-donor eyes. We were able to visualize at video-rate speed single RPE lipofuscin granules, demonstrating the potential to develop this method toward clinical practice for patients with RPE-related retinal disease like AMD.

We describe a novel scanning laser ophthalmoscope (SLO)-based on a video-rate second harmonic generation imaging microscope. A titanium:sapphire femtosecond laser was coupled to a modified SLO. The laser beam was scanned over the sample, and the light produced by second harmonic generation (SHG) was collected for imaging at video-speed. The device was used for imaging the lamina cribrosa (LC) of enucleated pig eyes. A resolution comparable to that of commercial multiphoton microscopes was reached. The SHG images were used for determining the average pore size of the LC determined from the images; the pressure dependence of the pore size was studied by the artificial increasing of the hydrostatic pressure in the eye. A pressure increase of 44.3 mmHg enlarged the average pore size of 62 analyzed pores by a statistically significant amount. The relative pore growth was measured at four different pressure levels in 25 pores. The pressure was increased in 15 mmHg steps. A general tendency for monothonic growth was observed, although single pores grew by no means linearly.

Background and Objective Diode pumped, all-solid-state ultrafast lasers are now widely used to perform minimally invasive refractive surgery and keratoplasty procedures. Despite such use, a systematic study concerning ultrafast laser–tissue interactions is lacking. We determined the corneal ablation threshold as a function of the laser pulse width and stromal depth by simultaneous monitoring of the intensity of the laser-induced plasma and the second harmonic generation signals (SHG) from the collagen. Study Design/Materials and Methods Ablation thresholds in porcine cornea samples were determined using three diode pumped all-solid-state ultrafast lasers (a Nd:glass femtosecond laser, a Yb:KYW femtosecond laser, and a Nd: YAG picosecond laser) over a range from 800 femtoseconds to 20 picoseconds. Results Corneal ablation threshold remained nearly constant within the first 200 µm of stroma and was consistent with previous findings with the threshold proportional to the square root of the laser pulse width. Conclusion Corneal ablation thresholds can be precisely determined by simultaneous monitoring of the intensity of the laser-induced plasma and the SHG from the cornea. Lasers Surg. Med. 39:654–658, 2007. © 2007 Wiley-Liss, Inc.

PURPOSE: To record the distribution and spectrum of human retinal pigment epithelial cell lipofuscin (LF) by two-photon-excited fluorescence (TPEF) and confocal laser scanning microscopy. METHODS: Ex vivo TPEF imaging of the human retinal pigment epithelium (RPE) of human donor eyes was conducted with a multiphoton laser scanning microscope that employs a femtosecond Ti:sapphire laser as an excitation laser source. The spectrum of autofluorescence of LF granules was analyzed with a confocal laser scanning microscope coupled to a UV argon laser. RESULTS: TPEF examination allowed for imaging of RPE cell morphology and intracellular distribution of LF granules with high-contrast and submicrometer resolution. Variations in cell size and shape as well as in autofluorescence spectra of individual LF granules were recorded. The typical diameter of LF granules was found to be below 1 mum, with some RPE cells possessing larger granules. Remarkably, enhanced blue-green autofluorescence was observed from these larger LF granules. CONCLUSIONS: TPEF imaging represents a novel tool for the investigation of morphologic and spectral characteristics of human RPE cells. Spectral variations of individual LF granules may indicate differences in the complex molecular composition. Compared to conventional single-photon excited autofluorescence, TPEF with a tunable laser source allows for reduced photo damage and deeper sensing depth. It may help to elucidate further the pathophysiological role of LF accumulation as a common downstream pathogenetic pathway in retinal diseases. With the proof of principle from this ex vivo study, further work is now planned to evaluate the safety of TPEF RPE imaging in RPE cultures and animal models.

Nd:glass femtosecond laser is promising as next generation mini-invasive eye surgical laser, with the advantages of excellent beam quality, high surgical precision and minimized side effects. However, there are still many open questions concerning the precision, efficiency and collateral effects of femtosecond laser refractive surgery. By non-invasive microscopic imaging methods including confocal, multiphoton, second harmonic and atomic force microscopy, we successfully characterized the three dimensional corneal ultrastructure without applying fixation and slicing. Based on the intrinsic properties of collagen, second harmonic cornea imaging proved to be outstanding to analyze the outcome of femtosecond laser intrastromal ablations. Strong contrast and large sensing depth second harmonic image was obtained without fixation, sectioning or labelling. The three dimensional ultrastructure of porcine cornea after Nd:glass femtosecond laser intrastromal surgery was examined to evaluate the concepts of minimum-invasive all-optical refractive eye surgery. No thermal damages were recognized and the surgical outcome appeared highly predictable. Due to the similarities between the physical principals of nonlinear laser scanning microscopy and femtosecond laser ablations, a setup of the Nd:glass femtosecond laser system integrating both the surgery and probing functions was proposed.

Wavefront technology was originally developed for the improvement of star images in optical astronomy. In 1978, this technology was for the first time applied to ophthalmology. At the University of Heidelberg, Germany, a closed loop adaptive optical system was designed to compensate for optical aberrations of the human eye. In the meantime, superresolution in retinal imaging has been achieved, providing detailed information on photoreceptors, small blood vessels, and nerve fiber structure in the human eye in vivo. Recently, new wavefront technology has been developed for the assessment of the human eye and it is now possible to apply a custom treatment to the eye, using the excimer laser. An adaptive optical closed loop system has been devised for preoperative simulation of refractive outcomes of aberration-free refractive surgical procedures. Treatment of a patient’s entire wavefront error has been demonstrated to improve the patient’s visual acuity beyond best spectacle-corrected vision.

Diode pumped Nd:glass all-solid-state femtosecond laser is promising for next generation refractive surgery, with the advantages of excellent surgical precision, minimal tissue damage and improved system stability and compactness. The microscopic evaluation of the outcome of femtosecond laser surgery is crucial before clinical applications. By two-photon laser scanning microscopy and non-invasive second harmonic imaging, the three dimensional ultrastructure of the porcine cornea is visualized without requiring slicing or staining. The minimal-invasive corneal flap cutting and non-invasive intrastromal surgery are investigated. Femtosecond laser intrastromal surgery demonstrated high ablation precision and mimimal side effects. However, there are elongated filaments/streaks observed in the cornea stroma, most likely due to the focusing optics and self-focusing.

Refractive surgery in the pursuit of perfect vision (e.g. 20/10) requires firstly an exact measurement of abberations induced by the eye and then a sophisticated surgical approach. A recent extension of wavefront measurement techniques and adaptive optics to ophthalmology has quantitatively characterized the quality of the human eye. The next milestone towards perfect vision is developing a more efficient and precise laser scalpel and evaluating minimal-invasive laser surgery strategies. Femtosecond all-solid-state MOPA lasers based on passive modelocking and chirped pulse amplification are excellent candidates for eye surgery due to their stability, ultra-high intensity and compact tabletop size. Furthermore, taking into account the peak emission in the near IR and diffraction limited focusing abilities, surgical laser systems performing precise intrastromal incisions for corneal flap resection and intrastromal corneal reshaping promise significant improvement over today's Photorefractive Keratectomy (PRK) and Laser Assisted In Situ Keratomileusis (LASIK) techniques which utilize UV excimer lasers. Through dispersion control and optimized regenerative amplification, a compact femtosecond all-solid-state laser with pulsed energy well above LIOB threshold and kHz repetition rate is constructed. After applying a pulse sequence to the eye, the modified corneal morphology is investigated by high resolution microscopy (Multi Photon/SHG Confocal Microscope).

The optical errors caused by the elements of the optical apparatus of the eye, as the cornea or the lens, strongly degrade the image on the retina. These errors are lager for larger pupil sizes.

Measurements of the wavefront of light reflected from the retina of the human eye can be used to determine optical aberrations of the human eye for large pupils. An instrument based on the Hartmann-Shack principle was developed. The wavefront is refracted by a microlens array and detected by a CCD camera. In first clinical studies human volunteer eyes and preoperative and postoperative refractive surgical patient eyes have been examined. An adaptive optical closed loop system has been devised for preoperative simulation of refractive outcomes of aberration free refractive surgical procedures. [J Refract Surg 2000; 16: S608-S6101

We investigated refractive corneal surgery in vivo and in vitro by intrastromal photodisruption using a compact ultrafast femtosecond laser system. Ultrashort-pulsed lasers operating in the femtosecond time regime are associated with significantly smaller and deterministic threshold energies for photodisruption, as well as reduced shock waves and smaller cavitation bubbles than the nanosecond or picosecond lasers. Our reliable all-solid-state laser system was specifically designed for real world medical applications. By scanning the 5 micron focus spot of the laser below the corneal surface, the overlapping small ablation volumes of single pulses resulted in contiguous tissue cutting and vaporization. Pulse energies were typically in the order of a few microjoules. Combination of different scanning patterns enabled us to perform corneal flap cutting, femtosecond-LASIK, and femtosecond intrastromal keratectomy in porcine, rabbit, and primate eyes. The cuts proved to be highly precise and possessed superior dissection and surface quality. Preliminary studies show consistent refractive changes in the in vivo studies. We conclude that the technology is capable to perform a variety of corneal refractive procedures at high precision, offering advantages over current mechanical and laser devices and enabling entirely new approaches for refractive surgery.

The phase retardation of light induced by the birefringent parts of the human retina in vivo is measured with an electro-optical ellipsometer using the principle of confocal imaging. A scanning unit allows to examine an area of 25 degrees by 12.5 degrees on the retina with a resolution of 256 by 128 points with both, incident and exit beam transmitting the cornea at a fixed position. Due to this fixed beam position the Muellermatrix of the cornea can be calculated using light which is specularly reflected on blood vessels lying above the nerve fiber layer of the retina. The obtained images show a homogenous radial distribution of the retinal retardation around the fovea without the appearance of the so called Haidinger brushes. The areas with the thickest nerve fiber layers, the arcuate bundles, appear in their typical arc and were measured quantitatively. In addition, an alternative method for the compensation of corneal birefringence is evaluated by focusing the light beam onto the surface of the lens. Hereby, the measured area in the center of the cornea is 3 X 0.75 mm2.© (1996) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

The dependence of the fluence at the threshold of laser- induced optical breakdown on the laser pulse duration has been investigated experimentally and theoretically for human cornea, human enamel, and bovine brain tissue. For the experiments in the range from 100 fs to 200 ps, we used a femtosecond dye laser system and a picosecond Nd:YLF laser system. We observed a significant decrease of the fluence at the threshold when reducing the pulse duration. The measured dependence on the pulse duration is in good agrement with our model.© (1996) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

Three nonlinear (beta) -barium borate (BBO) crystals were used to generate the fourth and fifth harmonic of a Nd:YLF picosecond laser with a fundamental wavelength of 1053 nm. The UV laser pulses were used to study the surface ablation of bovine soft tissue. Whereas the general study was concentrated on brain tissue, first results were also obtained with cornea tissue. The ablation efficiency as a function of pulse energy and the ablation threshold were determined for both UV wavelengths. Histology and SEM- microscopy of the performed laser lesions are presented. Furthermore, time-resolved flash photographs we recorded by illuminating the ablation area with green picosecond laser pulses and detecting the probe beam with a CCD chip. The photographs yielded information about the physical properties of the ablation process. The emission of a shock wave into the air environment as well as the ejection of dissected ablation fragments was observed for pulse energies down to 10 (mu) J. Both UV wavelengths can be combined with an ophthalmic application system, which comprises an xyz- scanning unit for realizing precalculated ablation patterns and a z-tracker for detecting the current surface position of the cornea. The information of the z-tracker can be used to compensate the axial eye-movement with the xyz-scanning unit.© (1996) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

In order to predict corneal curvature change after intrastromal surgery three finite element models are developed and compared. The models differ essentially in the underlying assumptions about the mechanical behavior of the cornea. The first model assumes an isotropic corneal structure with a nonlinear stress/strain behavior, thus providing a lower limit for the corneal curvature change. Contrary model two assumes transverse isotropy combined with a linear stress/strain behavior, in order to provide an upper limit for corneal curvature change. Although differing in their structural assumptions, model one and two consider the cornea as being a solid structure. A third model, based on a multilayered thin shell theory, shows that this might not be sufficient. In addition the calculations are run for two different ablation patterns. The performed analyses are extremely useful in choosing ablation patterns and dimensions for intrastromal refractive surgery. But as the model predictions of corneal curvature change differ in about 4 dpt for a given pattern, more subtle modifications and a better knowledge of the mechanical parameters of the cornea are necessary in order to obtain applicability of model calculations to predict the actual outcome of a performed surgery.

Changes in parapapillary retina height may be an early indicator of glaucomatous tissue damage. The subject of this study is the development of an automatic computer analysis for the height variations of the parapapillary retina obtained from simultaneously taken pairs of stereophotographs of optic nerve heads. The exact evaluation of stereo photographs is, so far, processed manually in the USA (Takamoto, Boston) in a timeconsuming procedure. An automatic analysis can be considered as an essential improvement in glaucoma diagnosis.© (1994) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

Glaucomatous diseases are one of the main reasons for blindness in modern indu- stry nations. In its chronical form this disease leads to a reduction of the nerve fi-ber layer of the retina. Therefore it is a point of interest to measure the thickness of the nerve fiber layer in the human eye in vivo. Because of the birefringent abili- ties of the nerve fiber layer an ellipsometric method can be used to measure its thickness. The next sections will describe a method based on standard ellipsometry, but using electro-optical devices instead of mechanical ones to measure the nerve fiber layer thickness faster and with less errors then it is done by mechanical ellip- sometry. 1. Introduction An eye disease that is relatively wide spreaded in modern industry nations is the chronic Glaucoma that is caused by an abnormal high eye pressure. The average va- lue for the eye pressure lies by about 20 mbar and fluctuates about S mbar during the day. Glaucomatous diseases decrease the blood flow through the retina and da

A laser system is described that offers manifold applications in ophthalmic surgery. The Nd:YLF solid-state laser emits at a wavelength of 1.053 micrometers . With pulse durations of 40 psec less energy is required to obtain optical breakdown compared to nanosecond lasers. The lower pulse energy allows a substantial confinement of the laser-tissue interaction. With a highly sophisticated delivery system eye movements can be tracked during surgery and a precise placement of the laser focus can be achieved. The high transmission coefficient of ocular structures for wavelengths around 1 micrometers enables the Nd:YLF laser to perform several internal eye surgeries such as cataract fragmentation, intrastromal refractive surgery, internal sclerostomy, or iridotomy. First clinical results of some of these procedures are summarized.

Many problems in ophthalmology deal with the three-dimensional structure of various parts of the living human eye. A complete confocal laser scanning microscope for diagnosis of eye disorders and first clinical results are described. Geometric measurements of the cornea and topographic measurements of the retinal substructures are presented. In addition, laser scanning tomography can be used to assess quantitatively the morphometry of the retina of the human eye. The confocal imaging mode of the laser tomographic scanner (LTS) is applied to evaluate the thickness of the nerve fiber layer. The contrast of nerve fiber layer images can be enhanced by polarization-dependent imaging modes, such as Fourier ellipsometry (FE) and differential interference contrast (DIC) imaging. To improve the depth resolution, an active-optical focusing system is employed.

Degeneration of retinal pigment epithelial (RPE) cells severely impairs the visual function of retina photoreceptors. However, little is known about the events that trigger the death of RPE cells at the subcellular level. Two-photon excited autofluorescence (TPEF) imaging of RPE cells proves to be well suited to investigate both the morphological and the spectral characteristics of the human RPE cells. The dominant fluorophores of autofluorescence derive from lipofuscin (LF) granules that accumulate in the cytoplasm of the RPE cells with increasing age. Spectral TPEF imaging reveals the existence of abnormal LF granules with blue shifted autofluorescence in RPE cells of aging patients and brings new insights into the complicated composition of the LF granules. Based on a proposed two-photon laser scanning ophthalmoscope, TPEF imaging of the living retina may be valuable for diagnostic and pathological studies of age related eye diseases.

Nonlinear laser scanning microscopy is widely used for noninvasive imaging in cell biology and tissue physiology. However, multiphoton fluorescence imaging of dense, transparent connective tissue (e.g., cornea) is challenging since sophisticated labeling or slicing is necessary. High-resolution, high-contrast second harmonic generation (SHG) imaging of corneal tissue based on the intrinsic structure of collagen is discussed. The three-dimensional corneal ultrastructure in depths up to hundreds of microns can be probed noninvasively, without any staining or mechanical slicing. As an important application of second harmonic imaging in ophthalmology, the modification of corneal ultrastructure using femtosecond laser intrastromal ablation is systematically investigated to evaluate next-generation refractive surgical approaches.

To study the retinal surface in the human eye in normal and diseased states we used laser scanning tomography. The confocal arrangement of the laser tomographic scanner permits examination of retinal topography in the axis perpendicular to the retinal surface. The eyes examined with the laser tomographic scanner included normal eyes, eyes with macular holes, impending macular holes, radiation retinopathy, macular edema, photocoagulation scars, subfoveal scars, and serous detachment of the fovea associated with subretinal neovascularization. The laser tomographic scanner is a new method that allows measurements of the topography of the internal limiting membrane in the macular area and may improve our understanding of the pathophysiologic characteristics and treatment of a variety of disorders of the macula.

Laser scanning tomography can be used to quantitatively assess the morphometry of the retina of the human eye. In this paper, the confocal imaging mode of the Laser Tomographic Scanner (LTS) is applied to evaluate the thickness of the nerve fiber layer. The contrast of nerve fiber layer images can be enhanced by polarization dependent imaging modes, like Fourier - ellipsometry (F.E.) and differential interference contrast (DIC) imaging. To improve the depth resolution, an active-optical focusing system is employed.