PURPOSE To assess the efficacy, safety, and predictability of the VisuMax femtosecond laser (Carl Zeiss Meditec). METHODS In a prospective, multicenter clinical trial, 32 eyes of 17 patients underwent LASIK and flap creation with the VisuMax femtosecond laser and ablation with the MEL 80 excimer laser (Carl Zeiss Meditec). Mean patient age was 35 years (range: 23 to 52 years). Mean preoperative spherical equivalent refraction was -4.04 +/- 1.39 diopters (D)(range:-1.50 to -7.25 D), and the planned flap thickness was 110, 120, 140, and 150 microm. RESULTS At the 3-month postoperative examination, all eyes had best spectacle-corrected visual acuity of 20/25 and uncorrected visual acuity (UCVA) of 20/40; 91% of eyes reached UCVA of 20/20 or better. Ninety-four percent of eyes were within +/- 0.50 D of the planned correction. Complications were limited to one case of suction loss, and no other adverse events were noted at 3 months. The flaps were planar and the mean achieved central flap thickness was 118 +/-9.2 microm for an attempted thickness of 120 microm. CONCLUSIONS The investigators achieved LASIK outcomes exceeding the US Food and Drug Administration guidelines for refractive procedures in this series of myopic eyes using the VisuMax femtosecond laser and MEL 80 excimer laser. The curved contact lens of the applanation glass combined with low suction permitted continuous fixation during treatment.

We have studied the possible nonlinear effects on tissue and model substance with emphasis on self-focusing effect. The nature of this effect and its biological consequences are discussed. We have studied how the self-focusing action changes with applied laser power and its effect on various depths of the biological materials. Critical power for biological material is calculated, and the peak power required for surgical purposes is found to be greater than the critical power. Furthermore, we discuss other possible effects that self-focusing might have on refractive surgery.

PURPOSE To assess flap creation and stromal bed quality of 2 femtosecond refractive surgery lasers in laser in situ keratomileusis. SETTING Augenklinik am Neumarkt, Cologne, Germany. METHODS Corneal flaps were created in 115 freshly enucleated porcine eyes using the 60 kHz IntraLase FS laser (Advanced Medical Optics) and a prototype model of the Femto LDV femtosecond laser (Ziemer Ophthalmic Systems AG). The parameters that were evaluated included actual versus intended thickness by subtraction pachymetry, cutting and total suction time, quality of flap edges, and smoothness of flap beds. Confocal microscopy (Atos PLmu [Altos GmbH]) was used to objectively determine the root mean square (RMS) of the surface roughness of the stromal bed. RESULTS Cutting time was 31 seconds for the 60 kHz IntraLase FS laser and 38 seconds for the Femto LDV laser. With both lasers, the standard deviation in achieved versus intended flap thickness was small (136 microm +/- 10 and 130 +/- 9 microm, respectively). Under micromorphologic examination, stromal bed quality was slightly better with the IntraLase. The RMS of bed roughness was 1.6 +/- 0.5 microm with the IntraLase and 2.0 +/- 0.4 microm with the Femto LDV. Neither laser showed significant thermal or mechanical damage in adjacent tissue layers of the stromal bed. The laser-induced bubble layer was more pronounced with the IntraLase. CONCLUSION The laser cuts of the IntraLase FS and Femto LDV femtosecond lasers were equally smooth and of excellent quality. The standard deviation of the flap thickness was small and equal in both systems.

Laser-induced damage is studied in the rat corneal epithelium and stroma using a combination of time-resolved imaging and biological assays. Cavitation bubble interactions with cells are visualized at a higher spatial resolution than previously reported. The shock wave is observed to propagate through the epithelium without cell displacement or deformation. Cavitation bubble expansion is damped in tissue with a reduction in maximum size in the range of 54 to 59%, as compared to 2-D and 3-D cultures. Bubble expansion on nanosecond timescales results in rupture of the epithelial sheet and severe compression of cell layers beyond the bubble rim. In the stroma, the dense collagen lamellae strongly damped bubble expansion, thus resulting in reduced damage. The acute biological response of this tissue to laser pulses is characterized by confocal fluorescence microscopy. A viability assay of the epithelium reveals that only cells around the immediate site of laser focus are killed, while cells seen to undergo large deformations remain alive. Actin morphology in cells facing this mechanical stress is unchanged. Collagen microstructure in the stroma as revealed by second-harmonic imaging around the ablation site shows minimal disruption. These cellular responses are also compared to in vitro 2-D and 3-D cell cultures.

PURPOSE: To determine the usefulness of optical coherence tomography (OCT) as a tool in guiding the femtosecond (fs) laser in the creation of a sub-Bowman keratomileusis (SBK) flap in human eyes. DESIGN: A nonrandomized case series. METHODS: In a private research laboratory setting, we performed an in vitro investigation on human autopsy eyes. Five human cadaver eyes, unsuitable for transplantation, underwent flap creation with a fs laser. The laser procedure was controlled in real-time with an OCT system (Thorlabs HL AG, Luebeck, Germany) to ensure that the cut was placed just underneath the Bowman layer. The fs laser worked at a repetition rate of 10 MHz with a single-pulse duration of < 400 fs (pulse energy in the nanoJoule range). As a control, all eyes underwent histologic dissection (toluidine blue) and were examined using light microscopy (LM). RESULTS: Video monitoring of the flap creation supported the feasibility of real-time OCT monitoring of the fs laser flap creation process. A clear distinction of the corneal epithelium was possible in all eyes. The Bowman membrane was not identified in all donor eyes at the given resolution of the OCT used in this study. Still, LM examination confirmed that the real-time monitoring assured a positioning of the cutting plane at minimum distance underneath the Bowman layer. CONCLUSION: This small laboratory test offers evidence that real-time OCT monitoring of creation of a SBK flap using a fs laser is possible, thus ensuring that the flap is created at the proper depth.

Despite the increasing use of infrared lasers in medical, industrial, and military settings, data on threshold radiant exposures and median effective dose (ED(50)) as they relate to laser-tissue interaction are limited. Our goals were to determine the ED(50) for single-pulse, 1540-nm laser exposures in ex vivo and in vitro rabbit corneal models and to characterize the histopathological changes associated with the laser-tissue interaction. An erbium-glass laser was used to deliver single, 1540-nm wavelength pulses to 27 ex vivo and 24 in vitro rabbit corneal models. The ex vivo model was exposed to single pulses of 0.8-ms duration and radiant energies ranging from 17.61 J/cm(2) to 42.26 J/cm(2). The in vitro corneal models were exposed to single pulses of 0.8 ms duration and had radiant exposures ranging from 14.87 to 29.72 J/cm(2). Tissue exposure sites were observed for presence of a lesion immediately post-exposure and 24 h after exposure. Histopathological evaluations of tissue exposure sites were conducted 24 h after exposure. The ED(50) was determined to be 21.24 J/cm(2) for the in vitro rabbit corneal models and 30.86 J/cm(2) for the ex vivo corneal models. Both the in vitro and ex vivo models displayed similar histopathological responses of tissue necrosis and epithelial cell proliferation.

PURPOSE To assess the response of the cornea to hydrogel intracorneal lens (ICL) insertion or laser in situ keratomileusis (LASIK) with IntraLase (IntraLase Corp.) at the cellular level. SETTING Department of Ophthalmology, University of Texas Southwestern Medical Center, Dallas, Texas, USA. METHODS Twenty patients (29 eyes) were evaluated by in vivo confocal microscopy 1 to 6 months postoperatively: 20 eyes had LASIK with flap creation by IntraLase, and 9 eyes had ICL insertion (8 following IntraLase). RESULTS For LASIK with IntraLase, keratocyte activation and/or interface haze was detected in 8 of 20 eyes. The remaining eyes had interface particles but no cell activation. Keratocyte activation was generally limited to a few cell layers adjacent to the interface. However, 2 patients exhibited multiple layers of activation and increased extracellular matrix (ECM) reflectivity (haze) surrounding the interface by confocal microscopy. Both patients also had clinical haze and photophobia. For ICLs, following insertion, 5 of 9 eyes had activated keratocytes adjacent to the implant surfaces. The largest amount of cell activation and ECM haze detected by confocal microscopy was in 2 patients with significant clinical haze. Structures with an epithelioid morphology were detected on some implant surfaces. Epithelial thickness was 33.3 microm +/- 2.3 (SD) in the ICL eyes and 49.2 +/- 6.5 microm in the LASIK with IntraLase eyes. CONCLUSIONS Both LASIK with IntraLase and ICL insertion following IntraLase induced keratocyte activation, which may underlie clinical observations of haze in some patients. Intracorneal lens implant also induced thinning of the overlying corneal epithelium.

PURPOSE Precision in corneal flap cutting is essential in LASIK surgery. Current mechanical microkeratomes have a very good performance record; however, in a few cases, complications can occur during the microkeratome pass and flap cut. Femtosecond lasers offer an alternative to the mechanical cut and can provide additional features regarding the flap morphology. In this study, we analyzed femtosecond laser flaps regarding their morphology, cut accuracy, and histopathology. METHODS Forty-five fresh porcine cadaveric eyes were prepared for femtosecond laser flap cutting with the Femtec femtosecond laser system (20/10 Perfect Vision, Heidelberg, Germany). The eyes were assigned to three different thickness groups, with 120-, 140-, or 180-microm cut depth, respectively. In addition, different flap diameters ranging from 8.0 to 9.5 mm and rim edge angulations between 60 degrees and 90 degrees were performed. After the cut, the eyes were examined under a microscope regarding accuracy and potential defects, and flap thickness and diameter were measured. In addition, flaps were prepared for further histopathologic examination. RESULTS All flap cuts were easily performed without any intraoperative complications. Flap thickness measurements revealed a median (in micrometers) of 110.5 (intended thickness 120), 142.5 (intended 140), and 180.0 (intended 180), respectively. The flap diameter for an intended size between 8.0 and 9.5 mm was within a range of +/-0.4 mm, the median at the maximum was 0.3 mm off. Histopathology revealed very low to almost no changes in the stromal structure of the cornea and correct hinge angulations. CONCLUSIONS LASIK flap cuts were easily performed without any complications. The accuracy and morphology were very precise and consistent. Histopathology revealed a smooth cut with hinge angulations, as expected.

Femtosecond mode-locked lasers are now being used routinely in multiphoton fluorescence and autofluorescence spectroscopy, are just beginning to be used in refractive surgery, and may be used in the future diagnosis of skin cancer. Pulses from these lasers induce non-linear effects in resultant tissue interactions. Using a modified confocal microscope with dispersion compensation and accurate measurements of beam diameter, a very low threshold was measured for photochemical oxidation in cultured cells. The measured threshold showed non-linear photo-oxidation at a peak irradiance and photon-flux density of 8.4x10(8) W cm-2 and 3.4x10(27) photons cm-2 s-1, respectively (90-fs pulse). The impact of these findings is significant to those using ultrashort lasers because they provide a tangible reference point (microscope-independent) for the generation of photo-oxidative stress in laser-exposed tissues, and because they highlight the importance of dispersion compensation in minimizing collateral tissue damage.

A 48-year-old woman had bilateral wavefront-guided laser in situ keratomileusis for myopia with IntraLase corneal flap creation. In the right eye the cavitation bubbles were observed not only in the interface between the flap and the corneal bed but also in the anterior chamber disappearing after 30 minutes. After the procedure, uncorrected visual acuity is 20/25 in both eyes; and specular microscopy shows normal hexagonal cells and density. Although no postoperative complications were observed in our case, further follow-up is needed to examine the long-term effect of this phenomenon of IntraLase.

Femtosecond (fs) laser-based cell surgery is typically done in two different regimes, at kHz or MHz repetition rate. Formation of reactive oxygen species (ROS) is an often predicted effect due to illumination with short laser pulses in biological tissue. We present our study on ROS formation in single cells in response to irradiation with fs laser pulses depending on the repetition rate while focusing into the cell nucleus. We observed a significant increase of ROS concentration directly after manipulation followed by a decrease in both regimes at kHz and MHz repetition rate. In addition, effects of consecutive exposures at MHz and kHz repetition rate and vice versa on ROS production were studied. Irradiation with a MHz pulse train followed by a kHz pulse train resulted in a significantly higher increase of ROS concentration than in the reversed case and often caused cell death. In the presence of the antioxidant ascorbic acid, accumulation of ROS and cell death were strongly reduced. Therefore, addition of antioxidants during fs laser-based cell surgery experiments could be advantageous in terms of suppressing photochemical damage to the cell.

Optoacoustic imaging was used for ophthalmic imaging, especially of the ciliary body region, which is of interest in the treatment of glaucoma. The different tissue structures below the sclera of porcine and rabbit eyes in vitro could be differentiated up to a depth of more than 1.5 mm. Based on the optoacoustic signals, two-dimensional tomographic images could be generated for visualization of this region in a B-scan mode. In addition, changes during the coagulation process could be measured in real time, allowing the development of online control mechanisms for cyclophotocoagulation, which is an important therapy of glaucoma.

PURPOSE:Thin-flap keratomileusis is a procedure that minimizes LASIK flap thickness to preserve both the corneal epithelium and the maximum residual stroma. This study investigated the usefulness of optical coherence tomography (OCT) as a tool in guiding the femtosecond laser in the creation of a thin flap in human eyes in a non-randomized case series. METHODS:In a private research laboratory, an in vitro investigation was performed on human autopsy eyes. Five human cadaver eyes, unsuitable for transplantation, underwent flap creation with a femtosecond laser. The laser procedure was controlled in real-time with an OCT system (Thorlabs HL AG) to ensure that the cut was placed just underneath Bowman's layer. The repetition rate of the femtosecond laser was 10 MHz with a single-pulse duration of <400 femtoseconds (pulse energy in the nJ range). As a control, all eyes underwent histological dissection and were examined using light microscopy. RESULTS:Video monitoring of the flap creation supported the feasibility of real-time OCT monitoring of the femtosecond laser flap creation process. A clear distinction of the corneal epithelium was possible in all eyes. Bowman's layer was not identified in all donor eyes at the given resolution of the OCT device used in this study. Light microscopy demonstrated flaps approximately 50-mum thick, confirming that the real-time monitoring assured a positioning of the cutting plane at minimum distance underneath Bowman's layer. CONCLUSIONS:This study of five human cadaver eyes shows that real-time OCT monitoring of the creation of thin-flaps in LASIK using a femtosecond laser is possible, thus ensuring that the flap is created at the desired depth.

PURPOSE To evaluate a new method for visualizing femtosecond laser pulse-induced microincisions inside crystalline lens tissue. SETTING Laser Zentrum Hannover e.V., Hannover, Germany. METHOD Lenses removed from porcine eyes were modified ex vivo by femtosecond laser pulses (wavelength 1040 nm, pulse duration 306 femtoseconds, pulse energy 1.0 to 2.5 microJ, repetition rate 100 kHz) to create defined planes at which lens fibers separate. The femtosecond laser pulses were delivered by a 3-dimension (3-D) scanning unit and transmitted by focusing optics (numerical aperture 0.18) into the lens tissue. Lens fiber orientation and femtosecond laser-induced microincisions were examined using a confocal laser scanning microscope (CLSM) based on a Rostock Cornea Module attached to a Heidelberg Retina Tomograph II. Optical sections were analyzed in 3-D using Amira software (version 4.1.1). RESULTS Normal lens fibers showed a parallel pattern with diameters between 3 microm and 9 microm, depending on scanning location. Microincision visualization showed different cutting effects depending on pulse energy of the femtosecond laser. The effects ranged from altered tissue-scattering properties with all fibers intact to definite fiber separation by a wide gap. Pulse energies that were too high or overlapped too tightly produced an incomplete cutting plane due to extensive microbubble generation. CONCLUSIONS The 3-D CLSM method permitted visualization and analysis of femtosecond laser pulse-induced microincisions inside crystalline lens tissue. Thus, 3-D CLSM may help optimize femtosecond laser-based procedures in the treatment of presbyopia.

The hearing performance with conventional hearing aids and cochlear implants is dramatically reduced in noisy environments and for sounds more complex than speech (e. g. music), partially due to the lack of localized sensorineural activation across different frequency regions with these devices. Laser light can be focused in a controlled manner and may provide more localized activation of the inner ear, the cochlea. We sought to assess whether visible light with parameters that could induce an optoacoustic effect (532 nm, 10-ns pulses) would activate the cochlea. Auditory brainstem responses (ABRs) were recorded preoperatively in anesthetized guinea pigs to confirm normal hearing. After opening the bulla, a 50-microm core-diameter optical fiber was positioned in the round window niche and directed toward the basilar membrane. Optically induced ABRs (OABRs), similar in shape to those of acoustic stimulation, were elicited with single pulses. The OABR peaks increased with energy level (0.6 to 23 microJ/pulse) and remained consistent even after 30 minutes of continuous stimulation at 13 microJ, indicating minimal or no stimulation-induced damage within the cochlea. Our findings demonstrate that visible light can effectively and reliably activate the cochlea without any apparent damage. Further studies are in progress to investigate the frequency-specific nature and mechanism of green light cochlear activation.

BACKGROUND According to the Helmholtz theory of accommodation the loss of accommodation amplitude is caused by the growing sclerosis of the crystalline lens, whereas the ciliary muscle and the lens capsule are mainly uneffected by age. A permanent treatment method for presbyopia which offers a dynamic accommodation ability is a recent field of study. The concept followed in this paper uses femtosecond laser pulses to potentially overcome the loss of deformation ability of the crystalline lens by creating gliding planes inside the lens tissue to improve its flexibility. METHODS The aim of the study is to show that the flexibility of human donor lenses can be increased by applying tightly focused near infrared femtosecond laser pulses into the lens tissue. Thereby the tissue is separated by the photodisruption effect. A certain pattern of gliding planes is cut inside the tissue of 41 human donor lenses and the deformation ability of the lenses are compared using the Fisher spinning test before and after laser treatment. RESULTS The laser treatment results in an increased deformation ability of the crystalline lens. The lens a-p thickness increases on average by 97 microm+/-14 microm after the treatment. The Fisher spinning test shows an increase of 16% in deformation ability of the lens at a rotational speed of 1620 rpm. CONCLUSION The creation of gliding planes with a fs laser inside the crystalline lens tissue can change the deformation ability of the lens. This might be an indication for a possible method to treat presbyopia in future.

Purinergic signalling in rat GFSHR-17 granulosa cells was characterised by Ca(2+)-imaging and perforated patch-clamp. We observed a resting intracellular Ca(2+)-concentration ([Ca(2+)](i)) of 100 nM and a membrane potential of -40 mV. This was consistent with high K(+)- and Cl(-) permeability and a high intracellular Cl(-) concentration of 40 mM. Application of ATP for 5-15 s every 3 min induced repeated [Ca(2+)](i) increases and a 30 mV hyperpolarization. The phospholipase C inhibitor U73122 or the IP(3)-receptor antagonist 2-aminoethoethyl diphenyl borate suppressed ATP responses. Further biochemical and pharmacological experiments revealed that ATP responses were related to stimulation of P2Y(2) and P2Y(4) receptors and that the [Ca(2+)](i) increase was a prerequisite for hyperpolarization. Inhibitors of Ca(2+)-activated channels or K(+) channels did not affect the ATP-evoked responses. Conversely, inhibitors of Cl(-) channels hyperpolarized cells to -70 mV and suppressed further ATP-evoked hyperpolarization. We propose that P2Y(2) and P2Y(4) receptors in granulosa cells modulate Cl(-) permeability by regulating Ca(2+)-release.

PURPOSE: To determine the usefulness of optical coherence tomography (OCT) as a tool in guiding the femtosecond (fs) laser in the creation of a sub-Bowman keratomileusis (SBK) flap in human eyes. DESIGN: A nonrandomized case series. METHODS: In a private research laboratory setting, we performed an in vitro investigation on human autopsy eyes. Five human cadaver eyes, unsuitable for transplantation, underwent flap creation with a fs laser. The laser procedure was controlled in real-time with an OCT system (Thorlabs HL AG, Luebeck, Germany) to ensure that the cut was placed just underneath the Bowman layer. The fs laser worked at a repetition rate of 10 MHz with a single-pulse duration of < 400 fs (pulse energy in the nanoJoule range). As a control, all eyes underwent histologic dissection (toluidine blue) and were examined using light microscopy (LM). RESULTS: Video monitoring of the flap creation supported the feasibility of real-time OCT monitoring of the fs laser flap creation process. A clear distinction of the corneal epithelium was possible in all eyes. The Bowman membrane was not identified in all donor eyes at the given resolution of the OCT used in this study. Still, LM examination confirmed that the real-time monitoring assured a positioning of the cutting plane at minimum distance underneath the Bowman layer. CONCLUSION: This small laboratory test offers evidence that real-time OCT monitoring of creation of a SBK flap using a fs laser is possible, thus ensuring that the flap is created at the proper depth.

The use of ultrashort laser pulses for microscopy has steadily increased over the past years. In this so-called multiphoton microscopy, laser pulses with pulse duration around 100 femtoseconds (fs) are used to excite fluorescence within the samples. Due to the high peak powers of fs lasers, the absorption mechanism of the laser light is based on nonlinear absorption. Therefore, the fluorescence signal is highly localized within the bulk of biological materials, similar to a confocal microscope. However, this nonlinear absorption mechanism can not only be used for imaging but for selective alteration of the material at the laser focus: The absorption can on one hand lead to the excitation of fluorescent molecules of fluorescently tagged cells by the simultaneous absorption of two or three photons or on the other hand, in case of higher order processes, to the creation of free-electron plasmas and, consequently, plasma-mediated ablation. Typical imaging powers are in the range of tens of milliwatts using 100-fs pulses at a repetition rate of 80-90 MHz, while pulse energies needed for ablation powers are as low as a few nanojoules when using high numerical aperture microscope objectives for focusing the laser radiation into the sample. Since the first demonstration of this technique, numerous applications of fs lasers have emerged within the field of cellular biology and microscopy. As the typical wavelengths of ultrashort laser systems lie in the near infrared between 800 and 1000 nm, high penetration depth can be achieved and can provide the possibility of imaging and manipulating the biological samples with one single laser system.

PURPOSE To determine through safety studies the tissue effects and potential cataractogenesis of laser modification of the crystalline lens (photophaco modulation). SETTING Laser Zentrum Hannover, Hannover, Germany. METHODS Six fresh porcine lenses and 6 living rabbit eyes (with the contralateral eye as a control) were radiated with a low-energy femtosecond laser to induce lens fiber disruption. After 3 months, the rabbit eyes were extracted and tested for light scatter and lens function and fixed for histology and ultrastructure. RESULTS After laser treatment, all lenses displayed a tightly packed array of intralenticular bubbles, which resolved with time. In the porcine eyes, the bubbles coalesced unless spacing of 9 mum or greater was applied at an energy of 2 microJ. In the rabbit eyes, an energy of 1 microJ and spacing of 10 microm was chosen for transcorneal delivery, showing minimum bubble coalescence. After 3 months, the rabbit lenses showed good transparency, with only 1 rabbit having cataract formation unrelated to the laser. Laser scanning studies show essentially identical values for the back focal length and sharpness of focus (variability of back focal length). Ultrastructurally, the rabbit eyes showed a 0.5 microm electron dense border layer with adjacent normal lens architecture. CONCLUSIONS Femtosecond laser photodisruption of the ocular lens yields a self-limited lesion with bubbles that resolve with time. In living animal eyes, no cataract formation was found with no loss of lens function or induced light scatter after 3 months. These results suggest that use of a low-energy femtosecond laser might be safe when modifying the lens for presbyopia correction.

ABSTRACT:: The introduction of the excimer laser for keratorefractive surgery in the 1990s permanently reshaped the treatment landscape for correcting refractive errors, such as myopia, hyperopia, and astigmatism. Until that point, these treatments had relied on less predictable techniques, such as radial keratotomy and automated lamellar keratectomy. In recent years, other new technologies, along with increased understanding of the basic science of refractive errors, higher-order aberrations, biomechanics, and the biology of corneal wound healing, have allowed for a reduction in the surgical complications of keratorefractive surgery. Novel technologies, such as eye tracking, anterior segment imaging, the femtosecond laser, and asphericity-optimized and wavefront-guided custom laser in situ keratomileusis, have assisted refractive surgeons in achieving greater predictability of their laser vision correction procedures. Understanding the cascade of events involved in the corneal wound healing process and examination of how corneal wound healing influences corneal biomechanics and optics are crucial to improve the efficacy and safety of laser vision correction.

Keratomileusis, brainchild of Jose I. Barraquer Moner, was conceived and developed as the first stromal sculpting method to correct refractive error in 1948. The word "keratomileusis" literally means "sculpting" of the "cornea." Barraquer's first procedures involved freezing a disc of anterior corneal tissue before removing stromal tissue with a lathe. Over the years, the procedure continued to develop, first through the Barraquer-Krumeich-Swinger non-freeze technique where tissue was removed from the underside of the disc by a second pass of the microkeratome. In-situ keratomileusis was later developed by passing the microkeratome a second time directly on the stromal bed. The procedure became known as automated lamellar keratoplasty with the invention of an automated microkeratome and was further refined by replacing the disc without sutures and later by stopping the microkeratome before the end of the pass to create a hinged flap, as first demonstrated in 1989. The history of the excimer laser dates back to 1900 and the quantum theory, eventually leading to the discovery that 193-nm ultraviolet excimer laser pulses could photoablate tissue without thermal damage. Ultrastructural and wound healing studies confirmed that large area ablation could be performed in the central cornea. This was described as photorefractive keratectomy in 1986 and the first sighted eyes were treated in 1988. An excimer laser was first used to sculpt from the stromal bed under a hinged flap created manually using a trephine and scalpel in 1988. The incorporation of a microkeratome in 1990 finally led to laser in situ keratomileusis-LASIK-as we know it today.

PURPOSE : To measure the refractive index (RI) of the human corneal stroma in vivo using an objective Abbé refractometer (VCH-1) and to determine if RI of the stroma is related to age. METHODS : VCH-1 was used to measure RI at the central anterior stroma immediately after lifting the flap in neophyte patients preselected for laser in situ keratomileusis. Surgical procedures continued as preplanned after measuring the RI, and in binocular cases, measurements were taken from the right eye only. Corneal flaps were created using a femtosecond laser system. RESULTS : Mean RI (±SD, range) and age (±SD, range) values were 1.373 (±0.006, 1.358-1.385) and 36.55 (±12.26, 18-74). A significant linear correlation was found between age and RI. Least squares regression lines equating RI with age (x, years) were of the form: RI = 1.36911 + 0.000096x (r =+0.195; n = 115; P = 0.037). Mean RI (± SD) in the 2 age groups separated by the median age (≤34 and ≥35) were 1.371 (±0.007; n = 57) and 1.374 (±0.005; n = 58). The difference was statistically significant (P = 0.018). CONCLUSIONS : RI of the anterior stroma in vivo tends to be increased in older patients when the corneal flap is created using a femtosecond laser device.

The availability of knockout mouse species provide a highly versatile platform for critically examining the corneal wound healing response. We aimed to develop and characterize the wound healing response in a mouse model of intrastromal femtosecond laser (FSL) keratotomy. An intrastromal lamellar dissection using a Visumax FSL was performed on 16 wild type mice (C57BL6). The energy level was optimized at 150nJ. The FSL was programmed to perform a lamellar dissection at 50 µM depth without sidecut. The flap was not lifted. Fellow eyes were used as controls. Slit lamp photography and confocal microscopy were performed immediately before the mice were sacrificed 4 h, 1, 3, and 7 days post surgery. Corneas were harvested for immunocytochemistry, transmission electron microscopy (TEM) and light microscopy (LM). Confocal microscopy showed an absence of keratocytes in the area immediately surrounding the dissection plane. The dissection plane and individual FSL plasma cavitation bubbles were clearly evident on TEM. There was evidence of Keratocyte cell death along the laser resection plane on TEM. LM revealed the dissection plane at a 20 µM depth, although not all epithelial cell layers were intact. Staining for monocytes using antibodies for CD11b (cluster of differentiation 11b) showed early migration at the peripheries at 4 h that increased at 24 h and became more central in treated corneas (p<0.001). Apoptotic cells were evident on TUNEL (terminal deoxynucleotidyl transferase dUTP nick end labeling) assay in the immediate ablation zone and were significantly raised at 4 and 24 h (p<0.001). Ki67 (Kiel 67 protein) positive proliferating keratocytes are evident at 3 days and increased significantly by 7 days (p<0.001). Minimal fibroblast (cluster of differentiation 90, CD90) transformation was seen at 1 week. No myofibroblasts were detected. We have demonstrated that FSL lamellar cuts can be effectively performed on mice and that this model exhibits typical signs of the corneal wound healing response. This model could provide a ubiquitous platform in which to study corneal wound healing responses in both wild type and knockout mice species. The ability to create such a lamellar pocket may be utilizzd for intrastromal drug delivery.

AIM To optimise interfaces of endothelial buttons created with femtosecond (FS) lasers. SETTING Department of Ophthalmology, Hôtel-Dieu Hospital, Paris, France. METHODS Forty-two corneas were divided into five groups of various cutting patterns and a control group of 100 μm laser in situ keratomileusis flap creation. A single path full lamellar cut (500 μm) was applied to groups 1 and 2. The same full lamellar cut was applied twice to groups 3 and 4. Two successive lamellar cuts were performed in group 5 (350 and 150 μm). 60 kHz and 150 kHz were used respectively in groups 1, 3, 5, 6 and 2, 4. In each group, different laser settings were tested to obtain the best interface quality while delivering minimal energy to the stroma. The quality of stromal interfaces from created endothelial lenticules was observed using a scanning electron microscope. RESULTS Stromal adherences persisted after both the single- and double-path procedure, creating central irregularities on the endothelial lenticule. Among all groups and settings tested, the double-layer pattern (group 5) with FS full lamellar cut parameters set for diameter (mm), depth (μm), energy (μJ) and spot size/step (μm) respectively on 9.0 mm, 350 μm, 2.1 μJ, 4:4 μm and 8.3 mm, 150 μm, 0.9 μJ, 4:4 μm created the smoothest interfaces with the best reproducibility. CONCLUSIONS Buttons for endothelial keratoplasty can be created with FS laser with a stromal interface quality comparable with that of refractive surgery.

PURPOSE To measure the refractive index (RI) of the human corneal stroma in vivo using an objective Abbé refractometer (VCH-1) and to determine if RI of the stroma is related to age. METHODS The VCH-1 was calibrated against a standard subjective Abbé refractometer using 7 randomly selected turbid semiliquid media. VCH-1 was used to measure RI at the central midstroma immediately after lifting the flap in neophyte patients preselected for laser-assisted in situ keratomileusis. Surgical procedures continued as preplanned after measuring the RI; in binocular cases, measurements were taken from the right eye only. Flaps were created using a mechanical microkeratome. RESULTS The VCH-1 was capable of measuring the RIs of all 7 turbid semiliquid media. The average RMS difference between RI estimates according to the VCH-1 and standard subjective Abbé refractometer was 0.003. The mean RI (± SD) of the stroma was 1.369 (± 0.008; range, 1.356-1.390), and the mean age (± SD) of the subjects was 33.7 years (± 8.86; range, 18-56 years). A significant linear correlation was found between age and RI. Least squares regression lines equating RI with age (x, years) was of the form: RI = 1.35711 + 0.00034x (r =+0.383, n = 36, P = 0.011). CONCLUSIONS RI of the stroma in vivo has a tendency to be increased in older patients when the stroma is resected using a mechanical microkeratome.

PURPOSE OF REVIEW The corneal flap laser in-situ keratomileusis (LASIK) is among the most important determinants in the successful outcome of the surgery. Femtosecond lasers have evolved over the last decade to all but replace the mechanical microkeratome as the preferred method to create these flaps. With improvements in femtosecond laser technology, there has been a reduction in the time taken for the cut and the quality of the stromal bed has improved. Improved predictability has led surgeons to explore the possibility of thin flap LASIK. RECENT FINDINGS Corneal flaps created with the femtosecond laser have been shown to be more predictable in depth and have a more desirable planar morphology. Corneal flaps created by the femtosecond laser can be customized according to depth, profile, morphology, and side-cut configuration. Changes in the angulation of the side cut, to reduce the incidence of epithelial ingrowth, have not been corroborated by clinical evidence as yet. Thin flap LASIK, also referred to as sub-Bowman's keratomileusis, has the advantage of preserving more stroma and potentially reducing the incidence of corneal ectasia but seems to be associated with an increased incidence of interface haze. SUMMARY This review examines the advantages of creating a flap with the femtosecond laser and the various configurations of these flaps. It also explores the advantages of varying the thickness and profile of femtosecond flaps.

A novel approach for applanation-free femtosecond refractive surgery with the help of a contact liquid layer is presented. A laboratory device for performing corneal procedures is described based on a femtosecond-laser system which has been tested and evaluated by processing ex vivo pig eyes. With its help, flap cuttings for different flap thicknesses were performed. The accomplished corneal surfaces are comparable to already published results. The reproducibility of the flap thicknesses is very good, with a standard deviation of 10 µm. The processing and removal of an intrastromal lenticule as thin as 30 µm could be shown. The extraction of such lenticule through a corneal side channel could also be accomplished successfully and is a promising improvement of the overall surgery procedure.

The use of ultrashort pulse lasers is current in refractive surgery and has recently been extended to corneal grafting (keratoplasty). When performing keratoplasty, however, permanent degradation of the optical properties of the patient's cornea compromises the penetration depth of the laser and the quality of the incisions, therefore causing unwanted secondary effects. Additionally, corneal grafting needs considerably higher penetration depths than refractive surgery. Little data are available about the interaction processes of the femtosecond pulses in the volume of pathological corneas-i.e., in the presence of spherical aberrations and optical scattering. We investigate the influence of the focusing numerical aperture on the laser-tissue interaction. We point out that at low numerical apertures (NAs), tissue damage is produced below and above the focal region. We attribute this phenomenon to nonlinear self-focusing effects. On the other hand, at high NAs, spherical aberrations become significant when focusing at high depths for posterior surgeries, which also limit the cutting efficiency. As high NAs are advisable for reducing unwanted nonlinear effects and ensure accurate cutting, particular attention should be paid to aberration management when developing clinical femtosecond lasers.

Laser in Situ Keratomileusis (LASIK) is the most common surgical procedure for the correction of refractive deviations1. LASIK has been ongoing lately in our country, exclusively in private settings. This is the reason for which our clinic has no experience with LASIK procedure and complications following it.We are presenting a case of a 23 year-old female with traumatic flap displacement on the left eye, 5 years after the original LASIK operation. She was presented to our clinic on the 6(th) posttraumatic day. The patient was immediately operated and the flap repositioned. The case we present, is the only one treated to our clinic with such a vision-threatening complication of LASIK procedure.