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Snakes belonging to the genus Naja (Elapid family), also known as "spitting cobras", can spit venom towards the eyes of the predator as a defensive strategy, causing painful and potentially blinding ocular envenoming. Venom ophthalmia is characterized by pain, hyperemia, blepharitis, blepharospasm and corneal erosions. Elapid venom ophthalmia is not well documented and no specific treatment exists. Furthermore, accidental ejection of venom by non-spitting vipers, as Bothrops, also occurs. The Ex vivo Eye Irritation Test model (EVEIT) has enabled important progress in the knowledge of chemical ocular burns. Considering the lack of experimental animal model, we adapted the EVEIT to study venom ophthalmia mechanisms. Ex vivo rabbit corneas were exposed to venoms from spitting (Naja mossambica, Naja nigricollis) and non-spitting (Naja naja, Bothrops jararaca and Bothrops lanceolatus) snakes, and rinsed or not with water. The corneal thickness and the depth of damage were assessed using high-resolution optical coherence tomography (HR-OCT) imaging and histological analysis. All Naja venoms induced significant corneal edema, collagen structure disorganization and epithelial necrosis. Corneas envenomed by African N. mossambica and N. nigricollis venoms were completely opaque. Opacification was not observed in corneas treated with venoms from non-spitting snakes, such as the Asian cobra, N. naja, and the vipers, B. jararaca and B. lanceolatus. Moreover, Bothrops venoms were able to damage the epithelium and cause collagen structure disorganization, but not edema. Immediate water rinsing improved corneal status, though damage and edema could still be observed. In conclusion, the present study shows that the EVEIT model was successfully adapted to set a new experimental ex vivo animal model of ophthalmia, caused by snake venoms, which will enable to explore new therapies for venom ophthalmia.

Objectives Imaging techniques with high resolution are evolving rapidly for medical applications and may substitute invasive diagnostic techniques. The use of ultrahigh resolution optical coherence tomography (UHR-OCT) to image healthy and morphologically altered bladder tissue with virtual histology is evaluated ex vivo to define parameters necessary for future, diagnostically relevant in vivo systems. Here, special focus is on the visualization of the basement membrane zone. Methods Optical coherence tomography examinations were performed by using a modified commercial OCT system comprising a Ti:sapphire femtosecond laser to support an enhanced resolution of 3 μm axial × 10 μm lateral. Tomograms of 142 fresh human bladder tissue samples from cystectomies, radical prostatectomies, and transurethral tumor resections were recorded and referenced to histologic sections using standard hematoxylin and eosin staining. Results OCT of normal bladder mucosa allows for a clear differentiation of urothelium and lamina propria. The basement membrane zone is identified as a narrow, low-scattering band between these layers. This allows for reliable exclusion of invasion. Healthy urothelial tissue, carcinoma in situ, and transitional cell carcinoma can be differentiated using this imaging technique. Sensitivity of UHR-OCT for malignant bladder tissue could be determined to be 83.8%, and specificity to be 78.1%. Conclusions UHR-OCT is considered promising in the attempt to strive for fluorescence cystoscopy-guided virtual histology as a means of supporting therapeutic decisions for bladder neoplasia.

Ocular irritation testing is a common requirement for the classification, labelling and packaging of chemicals (substances and mixtures). The in vivo Draize rabbit eye test (OECD Test Guideline 405) is considered to be the regulatory reference method for the classification of chemicals according to their potential to induce eye injury. In the Draize test, chemicals are applied to rabbit eyes in vivo, and changes are monitored over time. If no damage is observed, the chemical is not categorised. Otherwise, the classification depends on the severity and reversibility of the damage. Alternative test methods have to be designed to match the classifications from the in vivo reference method. However, observation of damage reversibility is usually not possible in vitro. Within the present study, a new organotypic method based on rabbit corneas obtained from food production is demonstrated to close this gap. The Ex Vivo Eye Irritation Test (EVEIT) retains the full biochemical activity of the corneal epithelium, epithelial stem cells and endothelium. This permits the in-depth analysis of ocular chemical trauma beyond that achievable by using established in vitro methods. In particular, the EVEIT is the first test to permit the direct monitoring of recovery of all corneal layers after damage. To develop a prediction model for the EVEIT that is comparable to the GHS system, 37 reference chemicals were analysed. The experimental data were used to derive a three-level potency ranking of eye irritation and corrosion that best fits the GHS categorisation. In vivo data available in the literature were used for comparison. When compared with GHS classification predictions, the overall accuracy of the three-level potency ranking was 78%. The classification of chemicals as irritating versus non-irritating resulted in 96% sensitivity, 91% specificity and 95% accuracy.

In optical coherence tomography (OCT), differential contrast can be generated by resonant nanoparticles using spectral multiplexing. Differential contrast can be of interest for medical applications for improving detection specificity of structures with low endogenous contrast. Differential contrast has been shown using OCT systems with one bandwidth; how- ever, this requires post-processing that is time consum- ing and reduces image resolution. In this study, we used a dual-band OCT prototype system with two far separated bandwidths in the clinically relevant optical window, and in search for the optimal differential contrast-generating particles for this prototype system, three different gold nanorods (AuNR) samples were investigated. The samples with different particle volume, aspect ratio, and absorption-maximum were imaged in a highly scattering phantom and on chicken muscle. In vitro, differential contrast was observed for the nanorods large (NRL) sample having the absorption- maximum within one bandwidth of the OCT and an average length of 75 nm. For the smaller AuNR (48 nm length) with comparable absorption-maximum, the obtained signal intensities were too low for being visible, although differences in signal intensities be- tween both bandwidths could be measured. NRL optimal concentration for differential contrast using this prototype system is between 100 and 500 lg Au/mL (0.51-2.54 mM). These results demonstrate the potential of real-time imaging of differential contrast in dual-band OCT and motivate in vivo application of plasmon resonant AuNR in order to improve the detection sensitivity for structures that are difficult to identify by OCT such as small blood vessels.

In this study, the optical properties of two nano-sized polymer colloids in optical coherence tomography (OCT) were compared in vitro with respect to their potential use as contrast agents. We used two types of particles: compact hydrophobic spherical polystyrene (PS) particles and soft water-swollen nanogel (NG) particles both with grafted hydrophilic shell, both prepared at two different sizes (PS at 300 and 150 nm, NG at 300 and 200 nm). The OCT backscattering signals of the particles in a vessel-mimicking highly scattering agar/TiO2 phantom were compared on either number of particles or weight percent. Larger particles and higher concentrations produced higher OCT contrast. At each concentration tested, a markedly higher contrast was achieved by PS particles than NG particles. PS particles generated a markedly higher OCT contrast than the phantom at concentrations of at least 1 × 1010 or 0.1 % for PS 300 nm and at least 3 × 1011 particles/mL or 0.4 % for PS 150 nm. The contrast generated by NG 300 nm was above the phantom contrast at concentrations of at least 3 × 1011 particles/mL or 1 %, whereas NG 200 nm only at 4 %. At any given weight percent, the differences in OCT contrast between differently sized particles were much less evident than in the comparison based on particle number. PS 300 nm generated also a good contrast ex vivo on chicken muscle tissue. These results strongly suggest that PS spheres have strong potential as intravascular OCT contrast agent, while NG particles need further contrast enhancer for being used as OCT contrast agent.

Enhanced visualization of tissue contrast and morphological boundaries is demonstrated by analyzing OCT volume data in two distinct wavelength regions. The extension of this so-called simultaneous dual-band method to three dimensions is realized by a custom-built 3D spectral domain OCT system imaging in the 800 und 1200 nm wavelength domain. Color representations of the spectral differences of high resolution OCT volume data significantly simplify the discrimination of different tissue structures beyond the capabilities of cross-sectional spectroscopic OCT approaches.

In optical sampling, a transient waveform under study is sampled with time delayed laser pulses, achieving a high time resolution. The limitations in measurement time due to the mechanical adjustment of the time delay can be overcome by asynchronous optical sampling (ASOPS) [1], where two pulsed lasers with slightly detuned repetition rates are employed. This technique is commonly used for studying transient characteristics [1], for infrared spectrometry [2] as well as remote sensing applications [3] and optical coherence tomography [4]. However, a static difference in repetition rates leads to static scanning ranges, resulting in unnecessary large measurement times and large amounts of measured data especially for the latter applications.

Three dimensional optical coherence tomography (OCT) is introduced as a valuable tool to analyze the pathogenesis of corneal diseases. Here, OCT in combination with a novel in vitro model for the dry eye syndrome enables an improved understanding of the underlying damaging process of the ocular surface. En-face OCT projections indicate a deep structural damage of the epithelium and anterior stroma by osmotic forces.

Electronically controlled coherent linear optical sampling for low coherence interferometry (LCI) and optical coherence tomography (OCT) is demonstrated, using two turn-key commercial mode-locked fiber lasers with synchronized repetition rates. This novel technique prevents repetition rate limitations present in previous implementations based on asynchronous optical sampling. Adjustable scanning ranges and scanning rates are realized within an interferometric setup by full electronic control of the mutual time delay of the two laser pulse trains. We implement this novel linear optical sampling scheme with broad spectral bandwidths for LCI, optical filter characterization and OCT imaging in two and three dimensions.

Understanding of dry eye syndrome (DES) today is driven by in vivo analysis of tear osmolarity, tear film break up time, impression cytology and description of symptoms. Existing in vivo models of DES need severe alterations of tear production or corneal integrity. For a more detailed analysis of DES under particular environmental and treatment conditions a considerable lack of in vitro methods exists. The main disadvantage of current in vitro models is the limited experimental time frame of only several hours and the impossibility to evaluate healing of epithelial defects. In the present study, evidence is given that these restrictions can be overcome by modifying the established Ex Vivo Eye Irritation Test (EVEIT) to realize a model system for DES. This test is based on abattoir rabbit eyes allowing an experimental time frame of up to 21 days using self-healing corneal cultures. In first experiments it is demonstrated that different severity levels of dry eye can be simulated in the EVEIT system. High-resolution optical coherence tomography (OCT) is applied to monitor the initial phase of DES under evaporative stress acting on the cornea. We observed changes in corneal layer thicknesses and in scattering properties of the stroma, which are sensitive indicators of environmental stress leading to irritation of the ocular surface under dry eye conditions. The combination of corneal culture under desiccating conditions and OCT monitoring offers a new perspective in understanding and treating of DES and is expected to allow for significant pharmacological screening tests.

Understanding of dry eye syndrome (DES) today is driven by in vivo analysis of tear osmolarity, tear film break up time, impression cytology and description of symptoms. Existing in vivo models of DES need severe alterations of tear production or corneal integrity. For a more detailed analysis of DES under particular environmental and treatment conditions a considerable lack of in vitro methods exists. The main disadvantage of current in vitro models is the limited experimental time frame of only several hours and the impossibility to evaluate healing of epithelial defects. In the present study, evidence is given that these restrictions can be overcome by modifying the established Ex Vivo Eye Irritation Test (EVEIT) to realize a model system for DES. This test is based on abattoir rabbit eyes allowing an experimental time frame of up to 21 days using self-healing corneal cultures. In first experiments it is demonstrated that different severity levels of dry eye can be simulated in the EVEIT system. High-resolution optical coherence tomography (OCT) is applied to monitor the initial phase of DES under evaporative stress acting on the cornea. We observed changes in corneal layer thicknesses and in scattering properties of the stroma, which are sensitive indicators of environmental stress leading to irritation of the ocular surface under dry eye conditions. The combination of corneal culture under desiccating conditions and OCT monitoring offers a new perspective in understanding and treating of DES and is expected to allow for significant pharmacological screening tests.

The objective of this study was to compare optical coherence tomography (OCT) with conventional techniques such as KOH-preparation, culture and histology in the identification of the fungal elements in the nail. A total of 18 patients were examined; 10 with clinically evident onychomycosis in toe nails, two with psoriatic nail lesions, one with nail affection caused by lichen planus and five healthy controls. Serial in vivo OCT analyses of onychomycosis was performed prior to KOH-preparation, culture and punch biopsy of the nail plate for consecutive histology. Fungal elements were detected non-invasively in vivo using OCT in all 10 patients with histologically proven onychomycosis. Fungal elements were detectable as highly scattering elongated structures inside the nail plate, in the middle of the areas of homogeneous decrease in signal intensity. KOH-preparations and culture did reveal a positive result in 5/6 out of 10 patients. In patients with psoriasis, lichen planus as well as in the healthy controls, no fungal infection could be detected by either method used. OCT is a reliable, easy to use, non-invasive and non-destructive method to visualise fungal elements in vivo in onychomycosis, even in cases of false negative KOH-preparation and culture. Furthermore, OCT offers the opportunity to screen several areas of the same nail plate and to detect fungal elements during local or systemic therapy.

A fiber-based spectral domain optical coherence tomography (OCT) system is described, imaging simultaneously at 740 and 1300 nm central wavelengths. Real-time imaging is demonstrated with axial resolutions

We present a high-speed, high-resolution, non-mechanical time-domain method for low coherence interferometry, utilizing multiheterodyne detection via two mode-locked femtosecond lasers. Tomographic depth sensing over 150 mm with 5.9 kHz scanning rates and 8 µm depth resolution is demonstrated.

A time domain optical coherence tomography (OCT) system without moving parts is described, which is based on multiheterodyning utilizing two mode-locked femtosecond lasers. By synchronizing the two lasers to slightly different repetition rates and coupling to an interferometric OCT setup, we obtain amplitude-modulated beat signals representing the structure of the specimen under investigation. Our system is suitable for biological imaging as well as technical applications. We demonstrate high axial imaging depths of 150 mm with up to 5000 axial scans per second, achieving equivalent path scanning velocities of 750 m/s.

A Fourier domain optical coherence tomography system is demonstrated, employing a commercial supercontinuum light source for the 1300nm wavelength region. Ultrahigh axial resolution of 1mm and 100dB sensitivity are achieved.

An off-the-shelf turn-key supercontinuum light source based on a passively mode-locked fiber laser, a fiber amplifier and a highly nonlinear fiber is evaluated for its application in ultrahigh resolution optical coherence tomography (OCT). Two spectral bands — one red shifted and one blue shifted in reference to the wavelength of the fiber laser — are employed as low coherence sources in OCT. Usable spectral bandwidths exceed 300 nm centered either at 790 or 1340 nm, with corresponding coherence lengths of 1.2 and 2.6 —, respectively. Optimization of the spectrum for simultaneous imaging at both spectral bands results in spectral bandwidths exceeding 200 nm for each band after filtering. A free-space OCT setup supporting the full spectral bandwidth is introduced which allows for ultrahigh resolution OCT imaging using both spectral bands simultaneously. Axial free space resolutions were measured to be less than 2 and 4 μm at 840 and 1230 nm, respectively. This approach combines ultrahigh resolution obtained at 840 nm with large penetration depths at 1230 nm. Frequency compounding of the OCT images can be used for speckle reduction while extracting spatially resolved spectroscopic features facilitates enhanced image contrast.

So far the study of chemical burns has lacked techniques to define penetration kinetics and the effects of decontamination within biological structures. In this study, we aim to demonstrate that high-resolution optical coherence tomography (HR-OCT) can close this gap. Rabbit corneas were exposed ex vivo to 2.5% hydrofluoric acid (HF) solution, and microstructural changes were monitored in the time domain by OCT imaging. HF application and penetration resulted in shrinkage of the corneal thickness, interpreted as a result of osmolar changes and of loss of water-binding capacity, and a substantial increase in OCT signal amplitudes. The effectiveness of different rinsing solutions on the chemical burn was also evaluated. With tap water and with 1% calcium gluconate, the deep corneal stroma remained clear until the end of the rinsing period but became opaque afterwards. With Hexafluorine 1 , the cornea remained clear for 60 min after rinsing ceased. We conclude that HR-OCT can assist in the clinical evaluation of an ex vivo eye irritation test, and that decontamination of an HF burn using Hexafluorine 1 is efficient.

For chemical burns a considerable lack of methods exists for defining penetration kinetics and effects of decontamination within biological structures. We demonstrate that time-resolved high-resolution optical coherence tomography can close this gap by monitoring changes in scattering properties and thicknesses of rabbit cornea ex vivo after topical application of different corrosives. Modifications in the corneal microstructure due to direct chemical interaction or changes in the hydration state as a result of osmotic imbalance compromise the corneal transparency. The associated increase in light scattering intensity within the cornea is observed with high spatial and temporal resolution. Parameters affecting the severity of pathophysiological damage like diffusion velocity, depth of penetration, resistance of barriers, and effectiveness of emergency treatment procedures are obtained. This study demonstrates the potential of high-resolution OCT for the visualization and direct non-invasive measurement of specific interaction of chemicals with the eye, exemplified on hydrofluoric acid burn.

Background: Three dimensional skin equivalents are widely used in dermatopharmacological and toxicological studies and as autologous transplants in wound healing. In pharmacology, there is tremendous need for monitoring the response of engineered skin equivalents to external treatment. Transplantation of skin equivalents for wound healing requires careful verification of their quality prior to transplantation. Optical coherence tomography (OCT) is a non-contact, non-destructive imaging technique for living tissues offering the potential to fulfill these needs. This work presents an analysis of OCT for high-resolution monitoring of skin equivalents at different stages during the culture process. Methods: We developed a high-resolution OCT imaging setup based on a commercially available OCT system. A broadband femtosecond laser light source replaces the original superluminescence diode. Tomograms of living skin equivalents were recorded with an axial resolution of 3 μm and correlated with histology and immunofluorescence images. Comparison with standard low-resolution OCT is presented to emphasize the advantages of high-resolution OCT for this application. Results: OCT is particularly able to distinguish between different layers of skin equivalents including stratum corneum, epidermal and dermal layer as well as the basement membrane zone. The high-resolution OCT scans correlate closely with two key benchmarks, histology and immunofluorescence imaging. Conclusions: This study clearly demonstrates the benefits of high-resolution OCT for identifying living tissue structure and morphology. Compared with the current gold standard histology, OCT offers non-destructive tissue imaging, enabling high-resolution evaluation of living tissue morphology and structure as it evolves.

Introduction La brulure chimique oculaire resulte de la destruction de constituents biochimiques des cellules des tissus vivants lors d’un contact avec un produit chimique irritant ou corrosif. Materiels et Methodes Des etudes in vitro ont ete realisees pour simuler une brulure oculaire par hydroxyde de sodium (NaOH). La diffusion de NaOH est simulee au travers une membrane semi-permeable de cellophane (30 g/cm 2 , 2.5 micrometres). Un volume de NaOH (3 gouttes ou 3/31 millilitre) est mis a la surface de la membrane et l’evolution du pH est mesuree dans un compartiment mimant la chambre anterieure, contenant le chlorure de sodium (6 ml, 420 mosmoles/kg). Le modele de membrane semi-permeable est egalement utilise pour simuler un rincage complet de l’exposition a NaOH 2N apres 20 secondes ou 1 minute d’exposition. La « chamber anterieure » (25 ml de chlorure de sodium 420 mosmoles/kg) est alors exposee via la membrane a 50 ml de soude 2N. Resultats Pour NaOH 0.1N, 0.2N, 1N, 2N et 5N, le pH final dans la « chamber anterieure », apres 10 minutes, est respectivement, de 8.61 et 9.91, 11.25, 11.95 et 12.34. Le rincage complet a montre une meilleure efficacite avec un lavage amphotere et hypertonique comparativement a l’eau. Pour 20 secondes de contact et apres 3 minutes de rincage, le pH externe etait respectivement de 9.12 pour la solution amphotere et 12.8 pour l’eau. Apres 45 minutes, le pH interne est de 9.25 avec le lavage amphotere et de 11.5 avec le rincage a l’eau. Pour une minute de contact, les courbes de pH sont semblables avec une diminution retardee de pH. Le pH interne est de 9.4 avec la solution amphotere et 11.85 avec l’eau. Discussion Ces resultats sont en accord avec les observations cliniques ainsi qu’avec les resultats de modeles ex vivo valides. Conclusion Une approche in vitro simple peut permettre d’evaluer precisement l’agressivite d’un produit chimique et de comparer de maniere reproductible l’efficacite de solutions de decontamination.

The use of high-resolution optical coherence tomography OCT to visualize penetration kinetics during the initial phase of chemical eye burns is evaluated. The changes in scattering properties and thickness of rabbit cornea ex vivo were monitored after topical application of different corrosives by time-resolved OCT imaging. Eye burn causes changes in the corneal microstructure due to chemical interaction or change in the hydration state as a result of osmotic imbalance. These changes compromise the corneal transparency. The associated increase in light scattering within the cornea is observed with high spatial and temporal resolution. Parameters affecting the severity of pathophysiological damage associated with chemical eye burns like diffusion velocity and depth of penetration are obtained. We demonstrate the potential of high-resolution OCT for the visual- ization and direct noninvasive measurement of specific interaction of chemicals with the eye. This work opens new horizons in clinical evaluation of chemical eye burns, eye irritation testing, and product testing for chemical and pharmacological products. © 2007 Society of

Ultra-high resolution optical coherence tomography (OCT) imaging is demonstrated simultaneously at 840 nm and 1230 nm central wavelength using an off-the-shelf turn-key supercontinuum light source. Spectral filtering of the light source emission results in a double peak spectrum with average powers exceeding 100 mW and bandwidths exceeding 200 nm for each wavelength band. A free-space OCT setup optimized to support both wavelengths in parallel is introduced. OCT imaging of biological tissue ex vivo and in vivo is demonstrated with axial resolutions measured to be