High-resolution in vivo imaging of regimes of laser damage to the primate retina.

Purpose. To investigate fundamental mechanisms of regimes of laser induced damage to the retina and the morphological changes associated with the damage response. Methods. Varying grades of photothermal, photochemical, and photomechanical retinal laser damage were produced in eyes of eight cynomolgus monkeys. An adaptive optics confocal scanning laser ophthalmoscope and spectral domain optical coherence tomographer were combined to simultaneously collect complementary in vivo images of retinal laser damage during and following exposure. Baseline color fundus photography was performed to complement high-resolution imaging. Monkeys were perfused with 10% buffered formalin and eyes were enucleated for histological analysis. Results. Laser energies for visible retinal damage in this study were consistent with previously reported damage thresholds. Lesions were identified in OCT images that were not visible in direct ophthalmoscopic examination or fundus photos. Unique diagnostic characteristics, specific to each damage regime, were identified and associated with shape and localization of lesions to specific retinal layers. Previously undocumented retinal healing response to blue continuous wave laser exposure was recorded through a novel experimental methodology. Conclusion. This study revealed increased sensitivity of lesion detection and improved specificity to the laser of origin utilizing high-resolution imaging when compared to traditional ophthalmic imaging techniques in the retina.

Trends in melanosome microcavitation thresholds for nanosecond pulse exposures in the near infrared.

Thresholds for microcavitation of bovine and porcine melanosomes were determined using nanosecond laser pulses in the near-infrared (1000 to 1319 nm) wavelength regime. Isolated melanosomes were irradiated by single pulses (10 or 50 ns) using a Q-switched Spectra Physics Nd:YAG laser coupled with an optical parametric oscillator (1000 to 1200 nm) or a continuum laser at 1319 nm. Time-resolved nanosecond strobe photography after the arrival of the irradiation beam allowed imaging of microcavitation events. Average fluence thresholds for microcavitation increased nonlinearly with increasing wavelength from ∼0.5 J/cm2 at 1000 nm to 2.6 J/cm2 at 1319 nm. Fluence thresholds were also measured for 10-ns pulses at 532 nm and found to be comparable to visible nanosecond pulse values published in previous reports. Calculated melanosome absorption coefficients decreased from 925 cm-1 at 1000 nm to 176 cm-1 at 1319 nm. This trend was found to be comparable to the decrease in retinal pigmented epithelial layer absorption coefficients reported over the same wavelength region. Estimated corneal total intraocular energy retinal damage threshold values were determined in order to compare to current and proposed maximum permissible exposure (MPE) safe levels. Results from this study support recently proposed changes to the MPE levels.

Porcine skin damage thresholds for 0.6 to 9.5 cm beam diameters from 1070-nm continuous-wave infrared laser radiation.

There is an increasing use of high-power fiber lasers in manufacturing and telecommunications industries operating in the infrared spectrum between 1000 and 2000 nm, which are advertised to provide as much as 10 kW continuous output power at 1070 nm. Safety standards have traditionally been based on experimental and modeling investigations with scant data available for these wavelengths. A series of studies using 1070-nm infrared lasers to determine the minimum visible lesion damage thresholds in skin using the Yucatan miniature pig (Sus scrofa domestica) for a range of beam diameters (0.6, 1.1, 1.9, 2.4, 4.7, and 9.5 cm) and a range of exposure durations (10 ms to 10 s) is presented. Experimental peak temperatures associated with each damage threshold were measured using thermal imaging. Peak temperatures at damage threshold for the 10-s exposures were ∼10°C lower than those at shorter exposures. The lowest and highest experimental minimum visible lesion damage thresholds were found to have peak radiant exposures of 19 and 432 J/cm2 for the beam diameter-exposure duration pairs of 2.4 cm, 25 ms and 0.6 cm, 10 s, respectively. Thresholds for beam diameters >2.5 cm had a weak to no effect on threshold radiant exposure levels for exposure times ≤0.25 s, but may have a larger effect on thresholds for exposures ≥10 s.

Infrared skin damage thresholds from 1319-nm continuous-wave laser exposures.

A series of experiments were conducted in vivo using Yucatan miniature pigs (Sus scrofa domestica) to determine thermal damage thresholds to the skin from 1319-nm continuous-wave Nd:YAG laser irradiation. Experiments employed exposure durations of 0.25, 1.0, 2.5, and 10 s and beam diameters of ∼0.6 and 1 cm. Thermal imagery data provided a time-dependent surface temperature response from the laser. A damage endpoint of fifty percent probability of a minimally visible effect was used to determine threshold for damage at 1 and 24 h postexposure. Predicted thermal response and damage thresholds are compared with a numerical model of optical-thermal interaction. Resultant trends with respect to exposure duration and beam diameter are compared with current standardized exposure limits for laser safety. Mathematical modeling agreed well with experimental data, predicting that though laser safety standards are sufficient for exposures <10 s, they may become less safe for very long exposures.

Effect of laser thermal injury on Langerhans cells in mouse and hairless guinea pig epidermis.

To examine the effect of laser thermal injury on Langerhans cells (LC) within the epidermis, the dorsal skin of mice and hairless guinea pigs was exposed to varying levels of laser irradiation using a thulium laser at a wavelength of 2.0 µm. At 6, 24 and 48 h post irradiation, animals were euthanized, skin samples prepared for histology and the epidermis obtained and stained by major histocompatibility complex-II staining (mice) or ATPase assay (hairless guinea pigs) for the enumeration of LC. Mouse skin exhibited histological evidence of thermal damage at 24 h post irradiation at even the lowest dose (0.14 W) and decreases in the numbers of epidermal LC were observed at all doses and decreases were proportional to dose. In contrast, hairless guinea pig skin only showed consistent histological evidence of thermal damage at the highest dose of irradiation (0.70 W) at 24 and 48 h post irradiation and exhibited a statistically significant decrease in numbers of epidermal LC only at this dose. Thus, epidermal LC depletion occurred in the skin of both mice and hairless guinea pigs in response to laser treatment and the magnitude of depletion directly correlated with the extent of thermal damage both within and between species.

Novel insight into cellulose supramolecular structure through ¹³C CP-MAS NMR spectroscopy and paramagnetic relaxation enhancement.

(13)C CP-MAS NMR and paramagnetic relaxation enhancement provide novel insight into the supramolecular structure of solid cellulose I and II. Separable NMR signals associated with crystalline interiors, solvent accessible and inaccessible surfaces, as well as non-crystalline material are assigned and confirmed. For the first time solvent accessibility is evidenced and monitored through (13)C T1 NMR relaxation enhancement in paramagnetic medium. Established NMR signal assignments for cellulose I have been confirmed. Existing cellulose II resonance attributions have been modified and extended. Novel spectral fitting routines for cellulose II allow for the reproducible quantification of separable signal contributions. Results from NMR line shape analyses are straightforwardly introduced into a model for cellulose II supramolecular structure.

In-vitro retinal model reveals a sharp transition between laser damage mechanisms.

We use laser damage thresholds in an in-vitro retinal model, and computational simulations to examine the laser exposure durations at which damage transitions from photothermal to photochemical at 413 nm. Our results indicate a dramatic shift in 1-h damage thresholds between exposure durations of 60 and 100 s. The trend in our in-vitro results is similar to a trend found in a recent study where retinal lesions were assessed 1-h post laser exposure in the rhesus eye Our data suggest that nonthermal mechanisms did not significantly contribute to cell death, even for exposures of 60 s. Knowledge of the transition point, and lack of concurrent thermal and nonthermal damage processes, are significant for those wishing to devise a comprehensive computational damage model.

Infrared skin damage thresholds from 1940-nm continuous-wave laser exposures.

A series of experiments are conducted in vivo using Yucatan mini-pigs (Sus scrofa domestica) to determine thermal damage thresholds to the skin from 1940-nm continuous-wave thulium fiber laser irradiation. Experiments employ exposure durations from 10 ms to 10 s and beam diameters of approximately 4.8 to 18 mm. Thermal imagery data provide a time-dependent surface temperature response from the laser. A damage endpoint of minimally visible effect is employed to determine threshold for damage at 1 and 24 h postexposure. Predicted thermal response and damage thresholds are compared with a numerical model of optical-thermal interaction. Results are compared with current exposure limits for laser safety. It is concluded that exposure limits should be based on data representative of large-beam exposures, where effects of radial diffusion are minimized for longer-duration damage thresholds.

Trends in retinal damage thresholds from 100-millisecond near-infrared laser radiation exposures: a study at 1,110, 1,130, 1,150, and 1,319 nm.

BACKGROUND AND OBJECTIVES: Retinal damage thresholds from 100-millisecond exposures to laser radiation for wavelengths between 1,100 and 1,350 nm have never previously been established. We sought to determine the retinal damage threshold for 100-millisecond exposures of near-infrared (NIR) laser radiation wavelengths at 1,110, 1,130, 1,150, and 1,319 nm. These data were then used to create trends for retinal damage thresholds over the 1,100-1,350 nm NIR region based upon linear absorption of laser radiation in ocular media and chromatic dispersion of the eye. MATERIALS AND METHODS: The paramacula and macula areas of the retina in Macaca mulatta (rhesus) subjects were exposed for 100 milliseconds to NIR laser radiation wavelengths using a Coherent OPO laser for 1,110, 1,130, and 1,150 nm and a Lee laser for 1,319 nm. Probit analysis was used to establish the estimated damage threshold in the retina for 50% of exposures (ED(50)). Using trends of transmitted energy to the retina, refractive error of the eye and linear absorption of the retina, a scaling factor (SF) method was created to fit the experimental data, predicting retinal damage thresholds over the 1,100-1,350 nm region. RESULTS: The experimental retinal damage threshold, ED(50), for 100-millisecond exposures for laser radiation wavelengths at 1,110, 1,130, and 1,319 nm were determined to be 193, 270, and 13,713 mW of power delivered to the cornea, respectively. The retinal damage threshold for the 1,150 nm wavelength was statistically undetermined due to laser-power limitations, but was achieved in one out of three subjects tested. CONCLUSION: The SF predicts the experimental 100- millisecond NIR ED(50) value for wavelengths between 1,100 and 1,350 nm.

In vitro model that approximates retinal damage threshold trends.

Without effective in vitro damage models, advances in our understanding of the physics and biology of laser-tissue interaction would be hampered due to cost and ethical limitations placed on the use of nonhuman primates. We extend our characterization of laser-induced cell death in an existing in vitro retinal model to include damage thresholds at 514 and 413 nm. The new data, when combined with data previously reported for 532 and 458 nm exposures, provide a sufficiently broad range of wavelengths and exposure durations (0.1 to 100 s) to make comparisons with minimum visible lesion (in vivo) data in the literature. Based on similarities between in vivo and in vitro action spectra and temporal action profiles, the cell culture model is found to respond to laser irradiation in a fundamentally similar fashion as the retina of the rhesus animal model. We further show that this response depends on the amount of intracellular melanin pigmentation.

Damage thresholds for cultured retinal pigment epithelial cells exposed to lasers at 532 nm and 458 nm.

The determination of safe exposure levels for lasers has come from damage assessment experiments in live animals, which typically involve correlating visually identifiable damage with laser dosimetry. Studying basic mechanisms of laser damage in animal retinal systems often requires tissue sampling (animal sacrifice), making justification and animal availability problematic. We determined laser damage thresholds in cultured monolayers of a human retinal pigment epithelial (RPE) cell line. By varying exposure duration and laser wavelength, we identified conditions leading to damage by presumed photochemical or thermal mechanisms. A comparison with literature values for ocular damage thresholds validates the in vitro model. The in vitro system described will facilitate molecular and cellular approaches for understanding laser-tissue interaction.

Visible lesion thresholds with pulse duration, spot size dependency, and model predictions for 1.54-microm, near-infrared laser pulses penetrating porcine skin.

Er:glass lasers have been in operation with both long pulses (hundreds of microseconds) and Q-switched pulses (50 to 100 ns) for more than 35 yr. The ocular hazards of this laser were reported early, and it was determined that damage to the eye from the 1.54-microm wavelength occurred mainly in the cornea where light from this wavelength is highly absorbed. Research on skin hazards has been reported only in the past few years because of limited pulse energies from these lasers. Currently, however, with pulse energies in the hundreds of joules, these lasers may be hazardous to the skin in addition to being eye hazards. We report our minimum visible lesion (MVL) threshold measurements for two different pulse durations and three different spot sizes for the 1.54-microm wavelength using porcine skin as an in vivo model. We also compare our measurements to results from our model, based on the heat transfer equation and the rate process equation. Our MVL-ED50 thresholds for the long pulse (600 micros) at 24 h postexposure were measured to be 20, 8.1, and 7.4 J cm(-2) for spot diameters of 0.7, 1.0, and 5 mm, respectively. Q-switched laser pulses of 31 ns had lower ED50 (estimated dose for a 50% probability of laser-induced damage) thresholds of 6.1 J cm(-2) for a 5-mm-diam, top-hat spatial profile laser pulse.

Accurate measure of laser irradiance threshold for near-infrared photo-oxidation with a modified confocal microscope.

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.

Simultaneous Exposure Using 532 and 860 nm lasers for visible lesion thresholds in the rhesus retina.

The growth of commercially available, simultaneous multi-wavelength laser systems has increased the likelihood of possible ocular hazard. For example, many systems utilize frequency multiplying methods to produce combinations of visible, near-infrared, and ultraviolet wavelengths. Unfortunately, very little data exists to substantiate the current methods for estimating hazards from simultaneous lasing. To properly assess the retinal hazards from these wavelengths, the retinal effects of 10-s laser irradiation from 532 and 860 nm were determined in non-human primates for four different relative dosage combinations of these wavelengths. This pair of wavelengths represents the typical problem of a visible-wavelength laser combined with an in-band, infrared wavelength that is not as well focused at the retina-a situation difficult to address. To add confidence to the experimental results obtained, a theoretical thermodynamic model was developed to predict the minimal damage threshold for simultaneous wavelengths at 1 h post exposure. The new model calculations and the data obtained are compared with results from one currently accepted method of predicting relative exposure limits from multi-wavelength systems. In addition, the current ANSI-Z136-2000 standard was used to compute the combined MPEs for comparison with measured visible lesion thresholds. A total of 12 eyes were exposed using four different ratios of power levels (532/860 power rations) to determine the contribution to the damage levels from each wavelength. The experimental data were analyzed using probit analysis at both 1-h and 24-h post exposure to determine the minimum-visible-lesion (MVL) thresholds at ED50 values, and these thresholds at 24 h varied from 5.6 mW to 17 mW total intraocular power.