Safe exposure times for slit-lamp fundus biomicroscopy with high plus lenses.

PURPOSE: The purpose of this study was to calculate the retinal irradiance and maximum permissible exposure time (MPE) using high plus fundus biomicroscopy lenses. METHODS: Four Volk handheld condensing lenses (+78 D, +90 D, Superfield NC, and Super 66) were tested with biomicroscopes from three manufacturers (Nikon, Topcon, and Zeiss) on both high and medium illumination. Using the conservation of radiance theorem, the retinal irradiance was calculated and. with guidelines from the American National Standard Institute (ANSI). the maximum permissible exposure time was determined. RESULTS: The range of MPE time across all lenses and biomicroscopes was from 23 seconds to 74 seconds on high illumination setting. The average MPE times were: for the +78 D, 36 seconds; the Superfield NC, 57 seconds; the Super 66, 32 seconds; and the +90 D, 52 seconds. CONCLUSION: Retinal irradiances and safe exposure times are useful guidelines in both the clinical and academic setting. Fundus biomicroscopy with non-contact high plus condensing lenses produced calculated retinal irradiances and MPE times similar to those of other commonly used ophthalmic equipment. Awareness of the maximum permissible exposure time increases the safety of this valuable technique.     

Endoillumination during vitrectomy and phototoxicity thresholds

AIM: To assess the retinal phototoxicity hazards of and to provide safety margins for endoillumination during vitrectomy. METHODS: The absolute power and spectral distribution from various light sources and filter combinations that are commercially available for vitreous surgery were measured. The maximal exposure times based on the ICNIRP safety guidelines for photochemical and thermal injury of the aphakic eye were calculated. Additionally, the effect of various measures that reduce the risk of phototoxicity was evaluated. RESULTS: Measurements of the spectrum and energy indicated that the ICNIRP safety guidelines for photochemical retinal damage are exceeded within 1 minute for nine out of 10 combinations tested. With an additional 475 nm long pass filter, light levels below 10 mW, and a distance from light probe to retina of at least 10 mm, the allowable exposure time can be increased up to 13 minutes. Thermal damage can be anticipated when the light probe touches the retina. CONCLUSION: Commercially available light sources for endoillumination during vitrectomy are not safe with respect to photochemical retinal damage. Even with maximal precautions macular phototoxic damage remains a factual danger during vitrectomy.     

Transpupillary thermotherapy for age-related macular degeneration: long-pulse photocoagulation, apoptosis, and heat shock proteins

OBJECTIVE: To provide a biophysical foundation for using transpupillary thermotherapy (TTT) to manage choroidal neovascularization in age-related macular degeneration (ARMD). METHODS: Retinal temperature rise in laser therapy is proportional to retinal irradiance (laser power/area) for a particular spot size, exposure duration, and wavelength. TTT is a low irradiance, large spot size, prolonged exposure (long-pulse), infrared laser photocoagulation protocol. Results from an experimentally confirmed, finite element model of retinal light absorption and heat conduction are used to analyze laser parameter selection and its consequences. Results from apoptosis, heat shock protein and hyperthermia research are used to examine how chorioretinal damage from clinical procedures might be reduced. RESULTS: Chorioretinal thermal equilibration occurs during long-pulse TTT photocoagulation. Retinal temperature increases are similar in the RPE where laser radiation absorption is significant and in the adjacent neural retina where there is negligible radiation absorption. For parameters used to treat occult choroidal neovascularization in lightly-pigmented fundi (800-mW, 810-nm, 3-mm retinal spot diameter, 60-sec exposure duration), the maximum chorioretinal temperature elevation is calculated to be roughly 10 degrees C, significantly lower than the 20 degrees C temperature elevations measured in threshold, conventional short-pulse retinal photocoagulation. CONCLUSIONS: To achieve a preselected temperature rise, TTT laser power must be increased or decreased in proportion to the diameter rather than the area of the laser spot. Clinical power settings should be adjusted for fundus pigmentation and media clarity because both of these factors affect absorbed retinal irradiance and thus retinal temperature rise. Noninvasive thermal dosimetry currently is unavailable for clinical retinal photocoagulation, but potential thermometric techniques include MRI, liposomal-encapsulated dyes, multispectral imaging or reflectometry, and subretinal or episcleral thermometry. TTT may be useful not only as independent therapy, but also as an adjunct to PDT, antiangiogenic drugs and ionizing radiation therapy in the management of neovascular ARMD. Low temperature, long-pulse photocoagulation is a potential strategy for decreasing neural retinal damage in subsequent TTT or short-pulse photocoagulation and perhaps even for treating glaucoma or retinal degenerations.     

A2e mediated phototoxic effects of endoilluminators

AIMS: To compare the theoretical retinal threshold time for endoilluminators and experimental phototoxic effect using A2e laden retinal pigment epithelial (RPE) cells. METHODS: The spectral irradiances of three types of 20 gauge and 25 gauge endoilluminators, currently commercially available from two manufacturers, were evaluated in conditions where the total beam spectral power was divided by the beam spot size at an estimated use distance of 5 mm from the retina. The retinal threshold time was calculated using the guidelines recommended by the International Commission on Non-Ionizing Radiation Protection. In vitro, A2e laden cells were evenly exposed to light for 30 minutes with a standard endoilluminator positioned 1 cm above the cells and the cell viability was assessed by WST-1 assay. RESULTS: The retinal threshold times were within 1 minute for all the endoilluminators tested. A significant decrease in the viability of A2e laden RPE cells was observed after they were exposed to light from two of the three 20 gauge endoilluminators. Cell viability was not affected by the exposure to 25 gauge endoilluminators under the same conditions. There was no correlation between the theoretical threshold times and experimental data. CONCLUSIONS: Light exposure during vitrectomy can induce photochemical damage to the retina. Although the A2e laden RPE model may not correctly mimic a clinical situation, this model may be useful to estimate the possible photochemical damage to RPE cells that could not be deduced by a theoretical retinal hazard model.     

Safety standards and measurement techniques for high intensity light sources

Any analysis of the retinal hazards of ophthalmic instrumentation based on criteria derived from current standards which implicitly assume large eye movements and smaller pupil sizes than are present in many diagnostic and therapeutic situations requires some care in application. Simple measurements of the optical power from a source with the calculation of the total retinal irradiance are not sufficient for hazard evaluation. Thorough and reliable hazard analysis requires knowledge not only of the spectral irradiance and the imaging conditions, but also some quantitative assessment of any pathological changes possibly producing unusual susceptibility in the patient's eyes.     

Sensitivity to retinal light damage and surgical blood oxygen levels

Increased oxygen levels decrease the threshold for photochemical retinal damage. We measured arterial oxygen levels in a group of ophthalmic surgical patients. As expected, levels exceeded unanesthetized measurements by one to two times. Based on experimental data, this could decrease the threshold for light-induced retinal damage during ophthalmic surgery by 40% to 50%. While the clinical implications of light-induced retinal damage in surgical eye patients are unclear, it is prudent to take steps to minimize light exposure during surgery.     

Review of the macaque model of light damage with implications for the use of ophthalmic instrumentation

The methods of evaluation of experimental light damage to monkey eyes include ophthalmoscopy. ERG, fluorescein angiography, and light and electron microscopy. Thresholds for damage caused by 15 min to 4 hr exposures at specific wavelengths from 457.9 nm–590 nm range from a retinal irradiance of 2 mW/cm²–80 mW/cm² or higher. Levels above 1 hr thresholds for monkey eyes are obtainable in human eyes with the usual slit lamp microscope and Hruby lens combination used in the clinic. Levels at or below the 15 min threshold are obtainable in normal human eyes with standard coaxial illumination from operating microscopes. The important factor in comparing light sources is the spectral irradiance produced on the retina rather than brightness.     

RPE damage thresholds and mechanisms for laser exposure in the microsecond-to-millisecond time regimen

PURPOSE: The retinal pigment epithelium (RPE) cells with their strongly absorbant melanosomes form the highest light-absorbing layer of the retina. It is well known that laser-induced retinal damage is caused by thermal denaturation at pulse durations longer than milliseconds and by microbubble formation around the melanosomes at pulses shorter than microseconds. The purpose of this work was to determine the pulse width when both effects merge. Therefore, the RPE damage threshold and mechanism of the damage at single laser pulses of 5-micros to 3-ms duration were investigated. METHODS: An argon laser beam (lambda 514 nm) was externally switched by an acousto-optic modulator to achieve pulses with constant power in the time range of 5 micros up to 3 ms. The pulses were applied to freshly prepared porcine RPE samples serving as a model system. After laser exposure RPE cell damage was proved by the cell-viability stain calceinAM. Microbubble formation was detected by acoustic techniques and by reflectometry. RESULTS: At a pulse duration of 5 micros, RPE cell damage was always associated with microbubble formation. At pulses of 50 micros, mostly thermal denaturation, but also microbubble formation, was detected. At the longer laser pulses (500 micros, 3 ms), RPE cell damage occurred without any microbubble appearance. CONCLUSIONS: At threshold irradiance, the transition time from thermal denaturation to thermomechanical damage of RPE cells is slightly below the laser pulse duration of 50 micros.     

Retinal damage produced by intraocular fiber optic light.

We exposed the maculas of owl monkey eyes to light from an intraocular fiber optic light source similar to that used for human pars plana vitrectomy. Retinal irradiance was calculated at 0.22 W/cm2. Eyes were exposed for time intervals ranging from 30 minutes to five minutes and were observed after light treatment by fundus photography and fluorescein angiography. Tissue was obtained for light and electron microscopy by animal killing at one hour, 24 hours, one week, and four weeks. Fundus lesions were seen ophthalmoscopically as early as five hours following 30 minutes of light exposure. Significant damage to the photoreceptor layer and less damage to the pigment epithelium was present by light and electron microscopy as early as one hour after 30 minutes of light exposure. By one month complete loss of photoreceptors with Müller cell junctions between inner retina and flattened abnormal retinal pigment epithelium cells was observed. Fluorescein angiography revealed significant staining of the pigment epithelium and outer retina 24 hours after 30 minutes of light exposure. No leakage from retinal vessels occurred. At one month following light treatment, transmission of choroidal fluorescein through window defects in the pigment epithelium was present with no retinal staining. The threshold for ophthalmoscopically visible fundus lesions in this study was 15 minutes of light exposure. Ten minutes of light treatment was the threshold for microscopic changes. Short light exposures damaged the outer retina and spared the pigment epithelium. Removing a substantial amount of the infrared light from our light source did not protect the retina from damage. Removal of light between 400 and 500 nm is probably more helpful in protecting the retina. Intermittent light exposure of the retina seemed as harmful as uninterrupted illumination for the same cumulative period of time. We speculate that the retinal damage caused by intraocular fiber optic light has primarily a photic mechanism. Damage to the retinal pigment epithelium may be secondary to outer retinal damage. The present levels of intraocular light used for human pars plana vitrectomy are probably safe in most instances. Lengthy preretinal membrane stripping procedures during vitrectomy, however, may pose a threat of light damage to the retina. This damage must be appreciated as continued efforts are made to produce brighter sources of intraocular light for human pars plana vitrectomy.     

Safety of UVA-riboflavin cross-linking of the cornea

PURPOSE: To study potential damage to ocular tissue during corneal collagen cross-linking (X-linking) by means of the riboflavin/UVA (370 nm) approach. METHODS: Comparison of the currently used technique with officially accepted guidelines regarding direct UV damage and the damage created by the induced free radicals (photochemical damage). RESULTS: The currently used UVA radiant exposure of 5.4 mJ/cm and the corresponding irradiance of 3 mW/cm2 is below the known damage thresholds of UVA for the corneal endothelium, lens, and retina. Regarding the photochemical damage caused by the free radicals, the damage thresholds for keratocytes and endothelial cells are 0.45 and 0.35 mW/cm, respectively. In a 400-microm-thick cornea saturated with riboflavin, the irradiance at the endothelial level was 0.18 mW/cm, which is a factor of 2 smaller than the damage threshold. CONCLUSIONS: After corneal X-linking, the stroma is depopulated of keratocytes approximately 300 microm deep. Repopulation of this area takes up to 6 months. As long as the cornea treated has a minimum thickness of 400 microm (as recommended), the corneal endothelium will not experience damage, nor will deeper structures such as lens and retina. The light source should provide a homogenous irradiance, avoiding hot spots.     

Photochemical damage of the retina

Visual perception occurs when radiation with a wavelength between 400 and 760 nm reaches the retina. The retina has evolved to capture photons efficiently and initiate visual transduction. The retina, however, is vulnerable to damage by light, a vulnerability that has long been recognized. Photochemical damage has been widely studied, because it can cause retinal damage within the intensity range of natural light. Photochemical lesions are primarily located in the outer layers at the central region of the retina. Two classes of photochemical damage have been recognized: Class I damage, which is characterized by the rhodopsin action spectrum, is believed to be mediated by visual pigments, with the primary lesions located in the photoreceptors; whereas Class II damage is generally confined to the retinal pigment epithelium. The action spectrum peaks in the short wavelength region, providing the basis for the concept of blue light hazard. Several factors can modify the susceptibility of the retina to photochemical damage. Photochemical mechanisms, in particular mechanisms that arise from illumination with blue light, are responsible for solar retinitis and for iatrogenic retinal insult from ophthalmological instruments. Further, blue light may play a role in the pathogenesis of age-related macular degeneration. Laboratory studies have suggested that photochemical damage includes oxidative events. Retinal cells die by apoptosis in response to photic injury, and the process of cell death is operated by diverse damaging mechanisms. Modern molecular biology techniques help to study in-depth the basic mechanism of photochemical damage of the retina and to develop strategies of neuroprotection.     

In vitro multispectral diffuse reflectance measurements of the porcine fundus

PURPOSE: To quantify the reflectance of the swine fundus as a function of illumination angle and wavelength using a novel technique of intravitreal illumination. METHODS: Enucleated swine eyes were illuminated with a scanning monochromator coupled to a fiber-optic probe placed in the vitreous at several locations. Intravitreal illumination was used to reduce the effects of extraneous reflections and scatter from the anterior structures of the eye, including glints from the internal limiting membrane and blood vessels. Intravitreal illumination provided different light paths and additional information to apply to retinal reflectance modeling. A 25-mm(2) region of the illuminated fundus was imaged while the angle of illumination was varied over a maximum range of 22 degrees . Multispectral images from areas free of large blood vessels were acquired. The diffusely reflected intensity was integrated over the pupil with a solid angle of 0.028 steradians. The spectral reflectance function was calculated for multiple illumination angles. RESULTS: Multispectral fundus image sets were obtained for two enucleated swine eyes by using intravitreal illumination. The fundus spectral reflectance function showed decreasing reflectance with increasing illumination angle, rapid changes in the 430- to 480-nm range, and a fairly consistent reflectance decrease for 480 to 700 nm. CONCLUSIONS: Variations in fundus spectral reflectance with change in the illumination angle were found to deviate from Lambertian behavior, varying from Lambertian by 5% across the spectrum in one sample and 20% in a second sample. Intravitreal illumination resulted in markedly decreased extraneous reflections.     

Fluorophotometric assessment of blood-retinal barrier function after white light exposure in the rabbit eye

Fluorophotometry was performed in 14 rabbits after exposure of one eye to white light with an energy insufficient to cause visible phototoxic retinal damage as determined by ophthalmoscopy and fundus photography. Fluorescence measurements in the vitreous were performed before and 1 hr after i.v. injection of fluorescein. Ratios between the fluorescein concentrations in the exposed and in the non-exposed fellow eye were calculated after correction for the autofluorescence. The average ratio directly after light exposure had significantly increased (P = 0.005) as compared to pre-exposure values and was maximal one day after exposure (P less than 0.005). Four days after exposure the ratios had returned to pre-exposure values (P greater than 0.05). A significant linear correlation between age and the ratios directly after exposure was found (r = -0.67; P less than 0.01). Signs of phototoxic retinal damage were not found on ophthalmoscopy and fundus photography, nor on light and electron microscopic examination of the retinal pigment epithelium, neuro-retina or retinal capillaries 1 and 4 days after light exposure. A fluorophotometric assessment of the blood-retinal barrier (BRB) function after white light exposure appeared to be a more sensitive parameter of light-induced damage than morphological examination since light exposures at retinal irradiance levels below the threshold for ultrastructural damage resulted in a temporary BRB dysfunction that could be detected by fluorophotometry but not by the other methods.     

Retinal damage threshold of ophthalmic Q-switched Nd-YAG laser in monkey eyes

Q-Switched Nd-YAG laser was irradiated to the ocular fundus of 24 eyes of 12 monkeys (Macaca fascicularis) using MICRORUPTOR II (LASAG AG, Thun, Switzerland) through the Goldmann type three-mirror contact lens. Multimode irradiation conditions were used; the pulse duration of 12 nanoseconds, spot size of 80 micrometers, convergence angle of 16 degrees and with various energy levels from 0.05 to 2.8 mJ. After irradiation, the fundus changes were observed by ophthalmoscopy and color fundus photography at 24 hours, 48 hours, 1, 2 and 5 weeks, and by fluorescein angiography at 24 hours, 1 and 5 weeks. Immediately after the irradiation and at 2 and 5 weeks, eyeballs were enucleated and histological observations were carried out. The fundus changes were more evident at 24 hours than at the immediate period after irradiation. At 24 hours, findings by fundus photography and fluorescein angiography were compared: both findings were in agreement. Consequently the retinal damage threshold was determined from the 24-hour findings; the threshold in ED50 was 296.3 microJ with 95% confidence limits from 249.8 to 351.1 microJ. These values corresponded to the retinal energy density of 3.5 J cm-2. Histological observations revealed that the site of damage was mainly in the retinal pigment epithelium when the irradiation was below 1.4 mJ. When the energy was increased to 1.9 mJ, the tissue damage extended to the middle layer of the neuroretina and also disruption of the Bruch's membrane occurred, leading to subretinal hemorrhage. With the energy of 2.8 mJ, destruction extended over wide areas including whole layers of the choroid and retina, leading to vitreous hemorrhage. Over the period of 5 weeks, proliferation of the retinal pigment epithelial cells took place and replaced the damaged retinal tissues.     

Fluorescein angiography: Risks

Fluorescein angiography has become a very important diagnostic aid in ophthalmology. The principles, technique and interpretation are briefly discussed. The risks of fluorescein angiography include direct ones resulting from injection of the dye as well as possible retinal damage from light exposure during the procedure. Retinal irradiance levels from fluorescein angiography have been compared to retinal threshold levels in normal animal eyes and using conventional techniques, appear to be safe. Its safety, however, in patients with retinal or other ocular disorders is unknown and requires further investigation.     

周边视网膜变性广角多模影像研究进展

周边视网膜变性是眼科临床常见的病变,不同类型的变性影响不同的视网膜层次,可能对视力造成威胁。尽管现代眼底成像技术被应用于研究其病理生理机制,由于其所处的特殊部位,影像学检查困难,因此发病机制仍不清楚。本文总结了有关周边视网膜变性的多种广角成像技术的影像特征,包括超广角眼底成像、广角频域光学相干断层扫描、光学相干断层扫描血管成像、荧光素眼底血管造影等,及其发病机制或病理特点的新观点,为临床诊疗提供新的思路。由于样本量非常少,且缺乏前瞻性、长期的多模态影像的观察研究,因而目前仍无法全面评价不同类型病变的进展性及危险性。期望在更广的范围内应用多模态广角成像技术对此类疾病进行研究,指导临床干预决策。     

Vitreoretinal surgery with slit-lamp illumination combined with a wide-angle-viewing contact lens

PURPOSE: To evaluate the feasibility of illuminating the fundus by combining a wide-angle-viewing contact lens and slit lamp attached to a surgical microscope during retinal reattachment surgery and vitrectomy. DESIGN: An interventional study. METHODS: We combined slit-lamp illumination and a wide-angle-viewing contact lens to visualize the fundus during retinal reattachment surgery and vitrectomy. RESULTS: We clearly observed the fundus using this combination approach during conventional retinal reattachment surgery, which we completed without using an indirect ophthalmoscope. With slit-lamp illumination, the wide-angle-viewing contact lens provided a wider area of illumination than a planoconcave contact lens during vitrectomy in addition to an area of illumination as wide as that obtained using a light pipe. CONCLUSIONS: Combining slit-lamp illumination and a wide-angle-viewing contact lens seems to provide a useful alternative method for viewing the fundus during vitreoretinal surgery.     

Retinal oximetry using intravitreal illumination

PURPOSE: To demonstrate spectroscopic retinal oximetry measurements on arteries and veins in swine using intravitreal illumination. Retinal arterial and venous saturations are measured for a range of inspired O2 levels after pars plana vitrectomy. METHODS: Pars plana vitrectomy and intravitreal manipulations were performed on two female American Yorkshire domestic swine. Light from a scanning monochromator was coupled into a fiberoptic intraocular illuminator inserted into the vitreous. The retinal vessels were illuminated obliquely, minimizing vessel glints. Multispectral images of the retinal vasculature were obtained as the swine's arterial blood oxygen saturation was decreased from 100% to 67% in decrements of approximately 10%. Retinal vessel spectra were used to calculate oxygen saturation in selected arteries and veins. Arterial oxygen saturations were calibrated using blood gas analysis on blood drawn from a Swan-Ganz catheter placed in the femoral artery. RESULTS: Oblique illumination of retinal vessels using an intravitreal fiberoptic illuminator provided a substantial reduction in the central vessel glint usually seen in fundus images, thus simplifying the analysis of spectral data. The vessel shadows were displaced from the vessel image simplifying the light paths in the eye. Using a full spectral analysis simplified by the light path reductions, we calculated retinal vessel saturations. The reduction of glint allowed for increased accuracy in measuring retinal vessel spectral optical density. Abnormally low retinal venous oxygen saturations were observed shortly after pars plana vitrectomy. CONCLUSIONS: Retinal oximetry using intravitreal illumination has been demonstrated. As a research tool, intravitreal illumination addresses several difficulties encountered when performing retinal oximetry with transcorneal illumination.     

Selective targeting of the retinal pigment epithelium in rabbit eyes with a scanning laser beam

PURPOSE: Selective targeting of the retinal pigment epithelium (RPE) with repetitive laser pulses that minimize thermal damage to the adjacent photoreceptors is a promising new therapeutic modality for RPE-related retinal diseases. The selectivity of an alternative, more versatile scanning approach was examined in vivo by using a broad range of scanning parameters. METHODS: Acousto-optic deflectors repeatedly scanned the focus of a continuous wave (cw)-laser across the retina of Dutch belted rabbits, producing microsecond irradiation at each RPE cell. Two irradiation patterns forming separated lines (SEP) or interlaced lines (INT), different dwell times (2.5-75 micros), and repetition numbers (10 and 100 scans with 100-Hz repetition rate) were tested. Thresholds were evaluated by fundus imaging and angiography. Histology was performed for selected parameters. RESULTS: Selective RPE cell damage was obtained with moderate laser power. The angiographic threshold power decreased with pulse duration, number of exposures, and applying the INT pattern. Ophthalmoscopic thresholds, indicating onset of thermal coagulation, were higher than twice the angiographic threshold for most tested parameters. Histology confirmed selective RPE cell damage for SEP irradiation with 7.5 and 15 micros; slower scan speeds or closed lines caused photoreceptor damage. CONCLUSIONS: A cw-laser scanner can be set up as a highly compact and versatile device. Selective RPE damage is feasible with dwell times up to 15 micros. Greatest selectivity is achieved with short exposure times and separated scan lines. Interlaced lines and long exposure times facilitate heat conduction into photoreceptors. A scanner is an attractive alternative for pulsed selective targeting, because both selective targeting and thermal photocoagulation can be realized.     

Damage Thresholds for Exposure to NIR and Blue Lasers in an In Vitro RPE Cell System

PURPOSE: Until reliable nonanimal systems of analysis are available, animal models will be necessary for ocular laser hazard analysis and for evaluating clinical applications. The purpose of this work was to demonstrate the utility of an in vitro system for laser bioeffects by identifying photothermal and photochemical cytotoxicity thresholds for continuous-wave (cw) and mode-locked (ml) laser exposures. METHODS: Exogenous melanosomes were added to hTERT-RPE1 cells in exposure wells 1 day before laser exposure. Thermal or photochemical laser exposures were delivered to artificially pigmented retinal pigment epithelial (RPE) cultures, with subsequent assay for viability 1 hour after exposure. Beam delivery for the 1-hour photochemical exposures was via a modified culture incubator. The cytoprotective effect of pretreatment with two antioxidants was investigated. RESULTS: Phagocytosis of melanosomes by the RPE cells was efficient, yielding cultures of uniform pigmentation. The damage threshold for the thermal exposure was consistent with published in vivo results. Thresholds for both blue exposures (cw and ml) were identical. Overnight treatment of cells with ascorbic acid (AA) minimized cell death from both cw and ml blue laser exposure, whereas similar treatment with N-acetyl-L-cysteine (NAC) was less effective. CONCLUSIONS: The in vitro system described is suitable for measuring meaningful thermal and photochemical laser damage thresholds. The system is also useful in comparative laser bioeffects studies, such as comparisons between cw and ml laser exposures, cells with various degrees of pigmentation, and studies determining the efficacy and mechanisms of treatments altering the response of cells to lasers.