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Background and Objective: A novel computer-guided laser treatment for open-angle glaucoma, called patterned laser trabeculoplasty, and its preliminary clinical evaluation is described. Patients and Methods: Forty-seven eyes of 25 patients with open-angle glaucoma received 532-nm laser treatment with 100-μm spots. Power was titrated for trabecular meshwork blanching at 10 ms and sub-visible treatment was applied with 5-ms pulses. The arc patterns of 66 spots rotated automatically after each laser application so that the new pattern was applied at an untreated position. Results: Approximately 1,100 laser spots were placed per eye in 16 steps, covering 360° of trabecular meshwork. The intraocular pressure decreased from the pretreatment level of 21.9 ± 4.1 to 16.0 ± 2.3 mm Hg at 1 month (n = 41) and remained stable around 15.5 ± 2.7 mm Hg during 6 months of follow-up (n = 30). Conclusion: Patterned laser trabeculoplasty provides rapid, precise, and minimally traumatic (sub-visible) computer-guided treatment with exact abutment of the patterns, exhibiting a 24% reduction in intraocular pressure during 6 months of follow-up ( P < .01).

Transparent ocular tissues, such as the cornea and crystalline lens, can be ablated or dissected using short-pulse lasers. In refractive and cataract surgeries, the cornea, lens, and lens capsule can be cut by producing dielectric breakdown in the focus of a near-infrared (IR) femtosecond laser, which results in explosive vaporization of the interstitial water, causing mechanical rupture of the surrounding tissue. Here, we compare the texture of edges of lens capsule cut by femtosecond lasers with IR and ultraviolet (UV) wavelengths and explore differences in interactions of these lasers with biological molecules. Scanning electron microscopy indicates that a 400-nm laser is capable of producing very smooth cut edges compared to 800 or 1030 nm at a similar focusing angle. Using gel electrophoresis and liquid chromatography/mass spectrometry, we observe laser-induced nonlinear breakdown of proteins and polypeptides by 400-nm femtosecond pulses above and below the dielectric breakdown threshold. On the other hand, 800-nm femtosecond lasers do not produce significant dissociation even above the threshold of dielectric breakdown. However, despite this additional interaction of UV femtosecond laser with proteins, we determine that efficient cutting requires plasma-mediated bubble formation and that remarkably smooth edges are the result of reduced thresholds and smaller focal volume.

A recent study showed that 355-nm nanosecond lasers cut cornea with similar precision to infrared femtosecond lasers. However, use of ultraviolet wavelength requires precise assessment of ocular safety to determine the range of possible ophthalmic applications. In this study, the 355-nm nanosecond laser was evaluated for corneal and iris damage in rabbit, porcine, and human donor eyes as determined by minimum visible lesion (MVL) observation, live/dead staining of the endothelium, and apoptosis assay. Single-pulse damage to the iris was evaluated on porcine eyes using live/dead staining. In live rabbits, the cumulative median effective dose (ED50) for corneal damage was 231 J/cm2, as seen by lesion observation. Appearance of endothelial damage in live/dead staining or apoptosis occurred at higher radiant exposure of 287 J/cm2. On enucleated rabbit and porcine corneas, ED50 was 87 and 52 J/cm2, respectively, by MVL, and 241 and 160 J/cm2 for endothelial damage. In human eyes, ED50 for MVL was 110 J/cm2 and endothelial damage at 453 J/cm2. Single-pulse iris damage occurred at ED 50 of 208 mJ/cm2. These values determine the energy permitted for surgical patterns and can guide development of ophthalmic laser systems. Lower damage threshold in corneas of enucleated eyes versus live rabbits is noted for future safety evaluation.

Transparent ocular tissues such as cornea and crystalline lens can be precisely ablated or dissected using ultrafast ultraviolet, visible, and infrared lasers. In refractive or cataract surgery, cutting of the cornea, lens, and lens capsule is typically produced by dielectric breakdown in the focus of a short-pulse laser which results in explosive vaporization of the interstitial water and mechanically ruptures the surrounding tissue. Here, we report that tissue can also be disrupted below the threshold of bubble appearance using 400 nm femtosecond pulses with minimal mechanical damage. Using gel electrophoresis and liquid chromatography/mass spectrometry, we assessed photodissociation of proteins and polypeptides by 400 nm femtosecond pulses both below and above the cavitation bubble threshold. Negligible protein dissociation was observed with 800 nm femtosecond lasers even above the threshold of dielectric breakdown. Scanning electron microscopy of the cut edges in porcine lens capsule demonstrated that plasma-mediated cutting results in the formation of grooves. Below the cavitation bubble threshold, precise cutting could still be produced with 400 nm femtosecond pulses, possibly due to molecular photodissociation of the tissue structural proteins.

Background and Objectives: The therapeutic effect of laser treatment for macular diseases is related to the damage to the retinal pigment epithelium (RPE) and the subsequent restoration of the defect due to RPE proliferation. In contrast to conventional laser treatment, it is possible to damage the RPE selectively and to spare the photoreceptors by using repetitive microsecond laser pulses. It was the aim of the study to investigate the influence of pulse duration and number of pulses on angiographically and ophthalmoscopically visible retinal damage thresholds in order to optimize treatment modalities. Study Design/Materials and Methods: In total, 625 laser lesions with various parameters were applied to the retina in 11 eyes of 6 Chinchilla breed rabbits using an experimental laser system (Nd:YLF at 527 nm). Pulse duration (1.7 microseconds and 200 nanoseconds) and number of pulses (100, 10, and 1 pulses) were varied at a constant repetition rate of 100 Hz. Damage thresholds were determined in terms of ophthalmoscopic and fluorescein angiographic visibility, and the therapeutic window (TW; angiographic ED50 vs. ophthalmoscopic ED50) as well as the safety range (SR; angiographic ED84 vs. ophthalmoscopic ED16) between both thresholds were calculated. Selected laser lesions were evaluated by histology. Results: Generally, the ED50 radiant exposure for angiographic visibility decreases for shorter laser pulses and with an increase in the number of pulses. The TW for both pulse durations (1.7 microseconds and 200 nanoseconds) was wider with 100 pulses than with single pulses. The widest TW was found for 100 pulses at 200 nanoseconds pulse duration (5.9-fold above the angiographic threshold), and the smallest TW with a factor of 1.6 was found for 1.7 microseconds single pulses. In terms of SR, only irradiation with 100 pulses at 200 nanoseconds pulse duration was associated with a ratio > 2. Independently of pulse duration, histological examination of laser sites 1 hour after irradiation revealed widely intact photoreceptors, while the underlying RPE was damaged. Conclusions: Pulse duration and number of pulses have a significant influence on RPE damage thresholds and consecutively on TW and SR. Because fundus pigmentation in humans may vary intra- and interindividually by a factor of 2, a large TW and ideally also a large SR should be ensured in a clinical treatment context. In rabbits, the safety range with 200 nanoseconds pulses is higher than with the pulse duration of 1.7 microseconds currently in clinical use. These findings suggest the need for clinical pilot studies to prove whether these results can be transposed to the situation in humans. Lasers Surg. Med. 34:206–215, 2004. 2004 Wiley-Liss, Inc.

* BACKGROUND AND OBJECTIVE: In both clinical and animal studies, it has been shown that repetitive short laser pulses can cause selective retinal pigment epithelium damage (RPE) with sparing of photoreceptors. Our purpose was to determine the ophthalmoscopic and angiographic damage thresholds as a function of pulse durations by using different pulsed laser systems to optimize treatment modalities. * MATERIALS AND METHODS: Chinchilla breed rabbits were narcotized and placed in a special holding system. Laser lesions were applied using a commercial laser slit lamp, contact lens, and irradiation with a frequency-doubled Nd: YLF laser (wavelength: 527 nm; repetition rate: 500 Hz; number of pulses: 100; pulse duration: 5 µß, 1.7 µß, 200 ns) and an argon-ion laser (514 nm, 500 Hz, 100 pulses, 5 µß and 200 ms). In all eyes, spots with different energies were placed into the regio macularis with a diameter of 102 µp? (tophat profile). After treatment, fundus photography and fluorescein angiography were performed and radiant exposure for ED50 damage determined. Speckle measurements at the fiber tips were performed to determine intensity peaks in the beam profile. * RESULTS: Using the Nd:YLF laser system, the ophthalmoscopic ED50 threshold energies were 25.4 µ] (5 µß), 32 µ] (1.7 µß), and 30 µ] (200 ns). The angiographic ED50 thresholds were 13.4 µ] (5 µ8), 9.2 µ] (1.7 µß), and 6.7 µ] (200 ns). With the argon laser, the angiographic threshold for 5 µß pulses was 5.5 µ). The ophthalmoscopic threshold could not he determined because of a lack of power; however, it was >12 µ}. For 200 ms, the ED50 radiant exposures were 20.4 mW ophthalmoscopically and 19.2 mW angiographically. Speckle factors were found to be 1.225 for the Nd: YLF and 3.180 for the argon laser. Thus, the maximal ED50-threshold radiant exposures for the Nd:YLF were calculated to be 362 mj/cm2 (5 µß), 478 mj/cm2 (1.7 µ-s), and 438 mj/cm2 (200 ns) ophthalmoscopically. Angiographically, the thresholds were 189 mj/cm2 (5 µß), 143 mj/cm2 (1.7 µ-s), and 97 mj/cm2 (200 ns). For the argon laser, the maximal ED50 radiant exposure threshold was 170 mj/cm2 angiographically. * CONCLUSION: The gap between the angiographic and the ophthalmoscopic thresholds for the 200 ns regime (4.5 times above angiographic ED50) was wider than for the 1.7 µ$ regime (3.3 times above the angiographic ED50). This would suggest the appropriate treatment would be 200 ns pulses. However, histologies have yet to prove that nonvisible mechanical effects increase with shorter pulse durations and could reduce the "therapeutic window." When comparing the thresholds with 5 µß pulses from the argon and Nd:YLF laser, it demonstrates that intensity modulations in the beam profile must be considered. [Ophthalmic Surg Lasers 200233:400-409]

Application of femtosecond lasers to cataract surgery has added unprecedented precision and reproducibility but ocular safety limits for the procedure are not well-quantified. We present an analysis of safety during laser cataract surgery considering scanned patterns, reduced blood perfusion, and light scattering on residual bubbles formed during laser cutting. Experimental results for continuous-wave 1030 nm irradiation of the retina in rabbits are used to calibrate damage threshold temperatures and perfusion rate for our computational model of ocular heating. Using conservative estimates for each safety factor, we compute the limits of the laser settings for cataract surgery that optimize procedure speed within the limits of retinal safety.

Femtosecond lasers have added unprecedented precision and reproducibility to cataract surgery. However, retinal safety limits for the near-infrared lasers employed in surgery are not well quantified. We determined retinal injury thresholds for scanning patterns while considering the effects of reduced blood perfusion from rising intraocular pressure and retinal protection from light scattering on bubbles and tissue fragments produced by laser cutting. We measured retinal damage thresholds of a stationary, 1030-nm, continuous-wave laser with 2.6-mm retinal spot size for 10- and 100-s exposures in rabbits to be 1.35 W (1.26 to 1.42) and 0.78 W (0.73 to 0.83), respectively, and 1.08 W (0.96 to 1.11) and 0.36 W (0.33 to 0.41) when retinal perfusion is blocked. These thresholds were input into a computational model of ocular heating to calculate damage threshold temperatures. By requiring the tissue temperature to remain below the damage threshold temperatures determined in stationary beam experiments, one can calculate conservative damage thresholds for cataract surgery patterns. Light scattering on microbubbles and tissue fragments decreased the transmitted power by 88% within a 12 deg angle, adding a significant margin for retinal safety. These results can be used for assessment of the maximum permissible exposure during laser cataract surgery under various assumptions of blood perfusion, treatment duration, and scanning patterns.

Purpose To compare 2 optical patient interface designs used for femtosecond laser–assisted cataract surgery. Setting Optimedica Corp., Santa Clara, California, USA, and Centro Laser, Santo Domingo, Dominican Republic. Design Experimental and clinical studies. Methods Laser capsulotomy was performed during cataract surgery with a curved contact lens interface (CCL) or a liquid optical immersion interface (LOI). The presence of corneal folds, incomplete capsulotomy, subconjunctival hemorrhage, and eye movement during laser treatment were analyzed using video and optical coherence tomography. The induced rise of intraocular pressure (IOP) was measured in porcine and cadaver eyes. Results Corneal folds were identified in 70% of the CCL cohort; 63% of these had areas of incomplete capsulotomies beneath the corneal folds. No corneal folds or incomplete capsulotomies were identified in the LOI cohort. The mean eye movement during capsulotomy creation (1.5 sec) was 50 μm with a CCL and 20 μm with an LOI. The LOI cohort had 36% less subconjunctival hemorrhage than the CCL cohort. During suction, the mean IOP rise was 32.4 mm Hg ± 3.4 (SD) in the CCL group and 17.7 ± 2.1 mm Hg in the LOI group. Conclusions Curved contact interfaces create corneal folds that can lead to incomplete capsulotomy during laser cataract surgery. A liquid interface eliminated corneal folds, improved globe stability, reduced subconjunctival hemorrhage, and lowered IOP rise. Financial Disclosure Drs. Talamo, Culbertson, Batlle, Feliz, and Palanker are consultants to and Messrs. Gooding, Angeley, Schuele, Marcellino, and Andersen, and Ms. Essock-Burns are employees of Optimedica Corp., Sunnyvale, California, USA.

About a third of people in the developed world will undergo cataract surgery in their lifetime. Currently, cataract surgery is a manual procedure highly dependent on the surgical skills and complicating factors. We developed and tested an image-guided laser system to improve the precision and reproducibility of cataract surgery. A long-range Optical Coherence Tomography automatically discerns the anterior and posterior surfaces of the lens and cornea, and a co-registered femtosecond laser then performs capsulotomy, lens segmentation and corneal incisions. Capsular strength following laser capsulotomy and mechanical capsulorhexis were compared on cadaveric eyes, and retinal safety was verified on rabbits. 50 patients have undergone cataract surgery using the laser system. Eyes were examined ophthalmoscopically, and extracted capsules were analyzed using histology and SEM.

About one-third of people in the developed world will undergo cataract surgery in their lifetime. Although marked improvements in surgical technique have occurred since the development of the current approach to lens replacement in the late 1960s and early 1970s, some critical steps of the procedure can still only be executed with limited precision. Current practice requires manual formation of an opening in the anterior lens capsule, fragmentation and evacuation of the lens tissue with an ultrasound probe, and implantation of a plastic intraocular lens into the remaining capsular bag. The size, shape, and position of the anterior capsular opening (one of the most critical steps in the procedure) are controlled by freehand pulling and tearing of the capsular tissue. Here, we report a technique that improves the precision and reproducibility of cataract surgery by performing anterior capsulotomy, lens segmentation, and corneal incisions with a femtosecond laser. The placement of the cuts was determined by imaging the anterior segment of the eye with integrated optical coherence tomography. Femtosecond laser produced continuous anterior capsular incisions, which were twice as strong and more than five times as precise in size and shape than manual capsulorhexis. Lens segmentation and softening simplified its emulsification and removal, decreasing the perceived cataract hardness by two grades. Three-dimensional cutting of the cornea guided by diagnostic imaging creates multiplanar self-sealing incisions and allows exact placement of the limbal relaxing incisions, potentially increasing the safety and performance of cataract surgery.

PURPOSE: To evaluate the safety, selectivity, and healing of retinal lesions created using a continuous line scanning laser. METHODS: A 532-nm Nd:YAG laser (PASCAL) with retinal beam diameters of 40 μm and 66 μm was applied to 60 eyes of 30 Dutch-belted rabbits. Retinal exposure duration varied from 15 μs to 60 μs. Lesions were acutely assessed by ophthalmoscopy and fluorescein angiography. Retinal pigment epithelial (RPE) flatmounts were evaluated with live-dead fluorescent assay. Histological analysis was performed at 7 time points from 1 hour to 2 months. RESULTS: The ratios of the threshold of rupture and of ophthalmoscopic visibility to fluorescein angiography visibility (measures of safety and selectivity) increased with decreasing duration and beam diameter. Fluorescein angiography and live-dead fluorescent assay yielded similar thresholds of RPE damage. Above the ophthalmoscopic visibility threshold, histology showed focal RPE damage and photoreceptor loss at 1 day, without inner retinal effects. By 1 week, photoreceptor and RPE continuity was restored. By 1 month, photoreceptors appeared normal. CONCLUSION: : Retinal therapy with a fast scanning continuous laser achieves selective targeting of the RPE and, at higher power, of the photoreceptors without permanent scarring or inner retinal damage. Continuous scanning laser can treat large retinal areas within standard eye fixation time.

This study evaluates the effects of exposure duration, beam diameter, and power on the safety, selectivity, and healing of retinal lesions created using a continuous line scanning laser. A 532 nm laser (PASCALTM) with retinal beam diameters of 40 and 66 μm was applied to 60 eyes of 30 Dutch-Belted rabbits. Retinal exposure duration varied from 15 to 60 μs. Lesions were acutely assessed by ophthalmoscopy and fluorescein angiography (FA). RPE flatmounts were evaluated with live-dead fluorescent assay (LD). Histological analysis was performed at 1 hour, 1 and 3 days, 1 and 2 weeks, and 1 and 2 months following laser treatment. Ophthalmoscopic visibility (OV) of the lesions corresponded to photoreceptor damage on histological analysis at 1 hour. In subvisible lesions, FA and LD yielded similar thresholds of RPE damage. The ratios of the threshold of rupture and of OV to FA visibility (measures of safety and selectivity) increased with decreasing duration and beam diameter. Above the threshold of OV, histology showed focal RPE damage and photoreceptor loss at one day without inner retinal effects. By one week, continuity of photoreceptor and RPE layers was restored. By 1 month, photoreceptors appeared normal while hypertrophy and hyperpigmentation of the RPE persisted. Retinal therapy with a fast scanning continuous laser achieves selective targeting of the RPE and, at higher power, of the photoreceptors. The damage zone in the photoreceptor layer is quickly filled-in, likely due to photoreceptor migration from adjacent zones. Continuous scanning laser can treat large retinal areas within standard eye fixation time.

Background: It has been shown that selective retina treatment (SRT) using a train of 1.7 microseconds laser pulses allows selective damage of the retinal pigment epithelium (RPE) while sparing the adjacent photoreceptors and thus avoiding laser scotoma. It was the purpose of this work to investigate SRT laser effects with Q-switched pulses of only 8 nanoseconds in duration by evaluating the angiographic and ophthalmoscopic damage thresholds and the damage range by histology in a rabbit model. Materials and Methods: A flash lamp pumped frequency doubled (532 nm) Nd:YAG laser with 8 nanoseconds pulse duration was used. In total 210 laser lesions, each calculated to be 102 mm in diameter on retina, were applied through a slit lamp onto the fundus of six eyes of Chinchilla Bastard rabbits. The rabbits were irradiated with increasing energies with single pulses and a train of 10 laser pulses at 10 Hz. After treatment fundus photography and angiography were performed to determine the damage thresholds (ED50-probability of RPE cell damage and neurosensory retinal damage) as well as the safety range between both thresholds (ratio of angiographic ED86 vs. ophthalmoscopic ED14). Selected histology was taken for single and repetitive pulse lesions after treatment. Results: Angiographic and ophthalmoscopic ED50-thresholds decreased with increasing number of pulses. For single pulse application ophthalmoscopic and angiographic ED50 were determined to 365 and 144 mJ/cm 2 , respectively. Regarding 10 pulses 266 and 72 mJ/cm 2 were found. No retinal hemorrhages or disruptions were observed for both sets of parameters. The therapeutic window between angiographic and ophthalmoscopic threshold revealed a factor of 3.1 for single pulses and 2.3 for repetitive pulse irradiation. The safety range respectively had a factor of 0.8 (single pulses) and 1.7 (10 pulses). Histologic examination of laser lesions with single and repetitive pulses at radiant exposures within the therapeutic window—292 and 213 mJ/cm 2 respectively—revealed damaged RPE, intact Bruch’s membrane and choriocapillaries. Photoreceptors were partly spared but also damaged to various extents. Conclusions: Short laser pulses of 8 nanoseconds pulse duration can damage the RPE without retinal hemorrhage or disruption. Selective damage of the RPE without affecting the photoreceptors can only rarely be achieved due to the small safety range. Thus, so far microsecond laser pulses for SRT seems favorable compared to nanosecond pulses in order to prevent unintentional photoreceptor damage. Lasers Surg. Med. 40:20–27, 2008. 2008 Wiley-Liss, Inc.

Transpupillary thermo therapy (TTT) is a slow (60 seconds) photothermal treatment of the fundus with a near-infrared (780-810nm) laser irradiating a large spot (0.5- 1. mm) on the retina. Due to high variability in ocular tissue properties and the lack of immediately observable outcome of the therapy, a real-time dosimetry is highly desirable. We found that fundus spectroscopy and spectrally-resolved imaging allow for non-invasive real-time monitoring and dosimetry of TTT. A 795nm laser was applied in rabbit eyes for 60 seconds using a 0.86mm retinal spot diameter. The fundus was illuminated with a broadband polarized light, and its reflectance spectra were measured in parallel and cross-polarizations. The fundus was also imaged in selected spectral domains. At irradiances that do not create ophthalmoscopically visible lesions the fundus reflectance increases at the wavelengths corresponding to absorption of the oxygenated blood indicating the reduced concentration of blood in the choroid. Vasoconstrictive response of the choroidal and retinal vasculature during TTT was also directly observed using spectrally-resolved imaging. At irradiances that produce ophthalmoscopically visible lesions a rapid reduction of the fundus reflectance was observed within the first 5-10 seconds of the exposure even when the visible lesions developed only by the end of the 60 second exposure. No visible lesions were produced where the laser was terminated after detection of the reduced scattering but prior to appearance of the enhanced scattering.

Fast and noninvasive detection of cellular stress is extremely useful for fundamental research and practical applications in medicine and biology. We discovered that light scattering spectroscopy enables us to monitor the transformations in cellular organelles under thermal stress. At the temperatures triggering expression of heat shock proteins, the refractive index of mitochondria increase within 1 min after the onset of heating, indicating enhanced metabolic activity. At higher temperatures and longer exposures, the organelles increase in size. This technique provides an insight into metabolic processes within organelles larger than 50 nm without exogenous staining and opens doors for noninvasive real-time assessment of cellular stress.

Fast and non-invasive detection of cellular stress is useful for fundamental research and practical applications in medicine and biology. Using Light Scattering Spectroscopy we extract information about changes in refractive index and size of the cellular organelles. Particle sizes down to 50nm in diameter can be detected using light within the spectral range of 450-850 nm. We monitor the heat-induced sub-cellular structural changes in human RPE cells and, for comparison, in transfected NIH-3T3 cells which express luciferase linked to the heat shock protein (HSP). Using inverse light scattering fitting algorithm, we reconstruct the size distribution of the sub-micron organelles from the light scattering spectrum. The most significant (up to 70%) and rapid (20sec) temperature-related changes can be linked to an increase of refractive index of the 160nm sized mitochondria. The start of this effect coincides with the onset of HSP expression. This technique provides an insight into metabolic processes within organelles larger than 50nm without exogenous staining and opens doors for non-invasive real-time assessment of cellular stress, which can be used for monitoring of retinal laser treatments like transpupillary thermo therapy or PDT.

Introduction: The selective RPE treatment (SRT) is a new method, which targets retinal diseases associated with disorders in the retinal pigment epithelium (RPE). By applying a train of μs laser pulses, it is possible to selectively damage RPE cells while sparing the adjacent photoreceptors and the neural retina. Due to the ophthalmoscopic invisibility of the RPE effects we investigated an optoacoustic (OA) on-line dosimetry system to monitor RPE damage non-invasively. Material and Methods: For in vitro experiments porcine RPE was irradiated with a Nd:YLF laser pulse train (527nm, 1.7μs, 5-40μJ, 30 pulses, 100 Hz). Pressure waves (optoacoustic transients) generated at the RPE were measured with a piezoelectric transducer. The RPE cell damage was visualised by fluorescence microscopy by means of the vitality stain CalceinAM. During 27 patient treatments (527nm, 1.7μs, 50-150μJ, 30 pulses, 100 Hz) the optoacoustic signals were measured with an ultrasonic transducer embedded in the contact lens. The RPE leakage was visualized by fluorescein and ICG angiography. Results: In vitro: Below the RPE cell damage threshold, the optoacoustic transients from each single pulse are almost similar. With RPE damage, fluctuations of the individual transients are observed during the pulse train. These fluctuations can be explained by statistical irregular microbubble formation around the strong light absorbing melanosomes inside the RPE cells, which occur after the temperature exceeds the vaporization threshold. The transient microbubbles probably lead to RPE cell disruption. An optoacoustic value (OA-value) calculated from the fluctuations was defined in order to assess RPE damage. Patient treatment: If optoacoustic pulse-to-pulse fluctuations were measured, RPE leakage was observed in fluorescein and ICG angiography. In 96% of the irradiated areas, RPE-leakage in fluorescein angiography and OA-value correlated. The stronger the optoacoustic pulse-to-pulse fluctuations, thus the higher the OA-value, the more intense angiographic leakage was observed in ICG-angiography. Conclusion: A non-invasive optoacoustic on-line dosimetry control to monitor RPE damage during SRT was developed. In order to avoid invasive angiography, it is currently evaluated in a multicenter clinical SRT study.

The therapeutic effect of laser treatment for macular diseases is related to the damage to the retinal pigment epithelium (RPE) and the subsequent restoration of the defect due to RPE proliferation. In contrast to conventional laser treatment, it is possible to damage the RPE selectively and to spare the photoreceptors by using repetitive microsecond laser pulses. It was the aim of the study to investigate the influence of pulse duration and number of pulses on angiographically and ophthalmoscopically visible retinal damage thresholds in order to optimize treatment modalities.In total, 625 laser lesions with various parameters were applied to the retina in 11 eyes of 6 Chinchilla breed rabbits using an experimental laser system (Nd:YLF at 527 nm). Pulse duration (1.7 microseconds and 200 nanoseconds) and number of pulses (100, 10, and 1 pulses) were varied at a constant repetition rate of 100 Hz. Damage thresholds were determined in terms of ophthalmoscopic and fluorescein angiographic visibility, and the therapeutic window (TW; angiographic ED(50) vs. ophthalmoscopic ED(50)) as well as the safety range (SR; angiographic ED(84) vs. ophthalmoscopic ED(16)) between both thresholds were calculated. Selected laser lesions were evaluated by histology.Generally, the ED(50) radiant exposure for angiographic visibility decreases for shorter laser pulses and with an increase in the number of pulses. The TW for both pulse durations (1.7 microseconds and 200 nanoseconds) was wider with 100 pulses than with single pulses. The widest TW was found for 100 pulses at 200 nanoseconds pulse duration (5.9-fold above the angiographic threshold), and the smallest TW with a factor of 1.6 was found for 1.7 microseconds single pulses. In terms of SR, only irradiation with 100 pulses at 200 nanoseconds pulse duration was associated with a ratio2. Independently of pulse duration, histological examination of laser sites 1 hour after irradiation revealed widely intact photoreceptors, while the underlying RPE was damaged.Pulse duration and number of pulses have a significant influence on RPE damage thresholds and consecutively on TW and SR. Because fundus pigmentation in humans may vary intra- and interindividually by a factor of 2, a large TW and ideally also a large SR should be ensured in a clinical treatment context. In rabbits, the safety range with 200 nanoseconds pulses is higher than with the pulse duration of 1.7 microseconds currently in clinical use. These findings suggest the need for clinical pilot studies to prove whether these results can be transposed to the situation in humans.

Selective targeting the retinal pigment epithelium (RPE) while sparing adjacent tissue such as the photoreceptors hasbeen demonstrated by repetitively irradiating the fundus with a train of green µs-laser pulses. The aim of this study wasto investigate selective RPE effects alternatively by means of rapidly scanning a cw-laser beam across the RPE to obtainthe required µs-illumination times.The radiation of an Ar + laser (514 nm) was transmitted through a 25 µm core diameter fiber to a scanner unit. The fibertip was imaged onto the object plane with a magnification of 0.75. The beam was repetitively scanned across porcineRPE samples in vitro providing an irradiation time of 1.6 µs. Cell damage was investigated with a fluorescence viabilityassay.The ED 50 cell damage power was determined to 580 mW when applying 10 exposures with a repetition rate of 500 Hz.It decreases to 250 mW for 500 exposures and more. The threshold keeps constant for an axial focus displacement ofabout 250 µm.In conclusion, selective cell targeting has been proved with a laser scanning device. The technique can be adapted forclinical RPE treatment to a slit-lamp or by modifying a retina angiograph.KEY WORDS : fluorescence microscopy, melanosome, laser scanner, RPE, selective photocoagulation, viability assay

Laser photocoagulation is a well-established treatment modality for a variety of retinal disorders, but is difficult to use near the fovea due to thermal retinal destruction. Certain diseases, such as drusen maculopathy, are thought to be caused by a dysfunction of the Retinal Pigment Epithelium. For those diseases selective targeting of the RPE, sparing the adjoining photoreceptors, might be the appropriate treatment to avoid laser scotoma, as it has been shown with application of a train of ms laser pulses by Birngruber and Roider. Our new approach is to use a conventional green cw laser and rapidly scan a small laser spot over the retina so as to produce microsecond(s) -illumination at each RPE cell. Two scanning devices were developed using acousto-optic deflectors. For the in vitro experiments the ED50 value RPE cell damage was 170 mW with 100 exposures, scanning with a speed of 1 spot diameter/3 microsecond(s) . In vivo experiments demonstrated an angiographic ED50 threshold of 66 mW for 100 exposures while scanning with an effective illumination time of 5 microsecond(s) . The ophthalmoscopic threshold was higher than a factor of 2 times the angiographic ED50. Using separated scan lines we show selectivity in the form of surviving cells in between irradiated lines. Selective destruction of RPE cells is possible using laser-scanning devices.© (2002) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

In all laser treatments at the fundus of the eye the temperature increase is unknown. In order to optimize the treatment modalities, a noninvasive online temperature determination is preferable. Applying laser pulses to the fundus, thermoelastic stress waves are emitted based on the thermal expansion of the heated tissue, mainly the retinal pigment epithelium (RPE). The amplitude of the thermoelastic wave is proportional to the thermal expansion coefficient, which linearly depends on temperature between 30-80 degree(s)C for water. The method was evaluated for selective RPE- treatment in vitro and clinically using the microsecond(s) -laser pulses for treatment and temperature determination simultaneously. Conventional laser photocoagulation was investigated in vitro using an Ar Ion laser for coagulation and low-energy N2-pumped dye laser pulses to probe the temperature. In all cases, sufficient pressure amplitudes were detected either by a needle hydrophon in vitro or by a contact lens with embedded transducer during treatment. Depending on the treatment parameter, temperature increase of 60 degree(s)C were evaluated from the pressure transients. All temperatures detected are in close agreement to heat diffusion calculations. We demonstrated a noninvasive online method to detect retinal temperatures during laser treatments. This technique can be adapted to photocoagulation, PDT and TTT.© (2002) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.

Objective: The selective damage of the retinal pigment epithelium (RPE) with repetitive microsecond(s) laser pulses is a new technique for the treatment of several retinal diseases. RPE can selectively be damaged by simultaneously sparing off the adjacent photoreceptor tissue. Objective of this study is to investigate whether optoacoustic (OA) transients occurring during irradiation might be used to control the invisible treatment effect. Setup: A train of frequency doubled Nd:YLF laser pulses (527 nm, 1.7microsecond(s) pulse length, 500Hz rep. rate) were applied via a laser slit lamp on porcine RPE samples. The acoustic transients were recorded with a broadband transducer. Results: At low radiant exposures ( 150 mJ/cm2), the OA transients differ from pulse to pulse within a pulse train, which can be attributed to microbubble formation around the strong absorbing melanosomes inside the RPE cells. FFT spectra of the OA transients show slight differences in the frequency spectrum with the different radiant exposures.

The selective damage of the retinal pigment epithelium (RPE) is a new treatment method for several retinal diseases. By applying a train of microsecond(s) laser pulses it is possible to selectively damage these cells and simultaneously spare the adjacent photoreceptor and neural tissue. Due to the ophthalmologic invisibility of the RPE cell damage we investigate an optoacoustic (OA) control system to monitor the RPE cell damage. Setup: The irradiation was performed with a frequency doubled Nd:YLF laser by applying a train of +s laser pulses. In vitro, the OA transients were received by an ultrasonic broadband transducer. During treatment an OA contact lens with embedded transducer was used. In vitro: Freshly enucleated porcine RPE samples with CalceinAM as life/death staining were used. Below RPE cell damage threshold a classic thermoelastic transient was found. Above cell damage threshold the OA transient differs form pulse to pulse. This can be explained by microbubble formation around the strong absorbing melanosomes inside the RPE cells. In vivo: We found the same pulse to pulse deviations of the OA transient above the fluoresceine angiographic detectable RPE damage threshold during treatment. This system give us a new approach to non-invasively monitor the selective RPE treatment.

The selective microphotocoagulation is a new technique to damage the retinal pigment epithelium (RPE), which is desired for treatment of several retinal diseases. By applying a train of microsecond(s) laser pulses it is possible to selectively destroy these cells and simultaneously spare the adjoining photoreceptor and neural tissue. We applied microsecond laser pulses of a Nd:YLF laser (527 nm), at a repetition rate of 500 Hz to porcine RPE. The light is absorbed in the RPE and by thermoelastic expansion, an optoacoustic (OA) signal will be generated which could be measured by an ultrasonic transducer. With this setup, the baseline temperature increase at the RPE, during irradiation can be determined, since the optoacoustic pressure signal depends on the temperature of the irradiated RPE. We found a linear dependence of the OA amplitude to the RPE sample temperature. At higher irradiance we proved the formation of microbubbles and bubble collapse in the RPE with OA techniques.

Photothermal (PT) and Photoacoustic (PA) Step-Scan FT-IR measurements are successful tools for thickness determination of layered samples. Aiming to separate optically between different layered tissues, we have been able to distinguish skin, liver and muscle tissues by means of PT and PA measurement techniques. Performing Step-Scan FT-IR measurements, the PT and PA phase and amplitude signals follow the tissue's absorption peaks which corresponds to the transmission spectra. These results are supported and confirmed by a simulation using the Finite- Difference-Method whereat the basic spectral data from transmission measurements and some assumed thermal properties of investigated tissues have been inserted.

Decreasing the pulse duration helps confine damage, shorten treatment time, and minimize pain during retinal photocoagulation. However, the safe therapeutic window (TW), the ratio of threshold powers for thermomechanical rupture of Bruch's membrane and mild coagulation, also decreases with shorter exposures. Two potential approaches toward increasing TW are investigated: (a) decreasing the central irradiance of the laser beam and (b) temporally modulating the pulse. An annular beam with adjustable central irradiance was created by coupling a 532-nm laser into a 200-μm core multimode optical fiber at a 4-7 deg angle to normal incidence. Pulse shapes were optimized using a computational model, and a waveform generator was used to drive a PASCAL photocoagulator (532 nm), producing modulated laser pulses. Acute thresholds for mild coagulation and rupture were measured in Dutch-Belted rabbit in vivo with an annular beam (154-163 μm retinal diameter) and modulated pulse (132 μm, uniform irradiance "flat-top" beam) with 2-50 ms pulse durations. Thresholds with conventional constant-power pulse and a flat-top beam were also determined. Both annular beam and modulated pulse provided a 28% increase in TW at 10-ms duration, affording the same TW as 20-ms pulses with conventional parameters. C 2011 Society of Photo-Optical Instrumentation Engineers (SPIE). (DOI: 10.1117/1.3542045)

The selective retina treatment SRT targets retinal diseases associated with disorders in the retinal pigment epithelium RPE. Due to the ophthalmoscopic invisibility of the laser-induced RPE effects, we investigate a noninvasive optoacoustic real-time dosimetry system. In vitro porcine RPE is irradiated with a Nd:YLF laser 527 nm, 1.7-s pulse duration, 5t o 40J, 30 pulses, 100-Hz repetition rate. Generated acoustic transients are measured with a piezoelectric trans- ducer. During 27 patient treatments, the acoustic transients are mea- sured with a transducer embedded in an ophthalmic contact lens. After treatment, RPE damage is visualized by fluorescein angiographic leakage. Below the RPE damage threshold, the optoacoustic transients show no pulse-to-pulse fluctuations within a laser pulse train. Above threshold, fluctuations of the individual transients among each other are observed. If optoacoustic pulse-to-pulse fluctuations are present, RPE leakage is observed in fluorescein angiography. In 96% of the irradiated areas, RPE leakage correlated with the optoacoustic defined threshold value. A noninvasive optoacoustic real-time dosimetry for SRT is developed and proved in vitro and during patient treatment. It detects the ophthalmoscopically invisible laser-induced damage of RPE cells and overcomes practical limitations of SRT for use in private practice. © 2005 Society of Photo-Optical Instrumentation Engineers.