Photodynamic action of merocyanine 540 on Staphylococcus epidermidis biofilms

Photodynamic treatment (PDT) has been proposed as a new approach for inactivation of biofilms associated with medical devices that are resistant to chemical additives or biocides. In this study, we evaluated the antimicrobial activity of merocyanine 540 (MC 540), a photosensitizing dye that is used for purging malignant cells from autologous bone marrow grafts, against Staphylococcus epidermidis biofilms. Effect of the combined photodynamic action of MC 540 and 532 nm laser was investigated on the viability and structure of biofilms of two Staphylococcus epidermidis strains, RP62A and 1457. Significant inactivation of cells was observed when biofilms were exposed to MC 540 and laser simultaneously. The effect was found to be light dose-dependent but S. epidermidis 1457 biofilm proved to be slightly more susceptible than S. epidermidis RP62A biofilm. Furthermore, significant killing of both types of cells was attained even when a fixed light dose was delivered to the biofilms. Confocal laser scanning microscope (CLSM) analysis indicated damage to bacterial cell membranes in photodynamically treated biofilms, while disruption of PDT-treated biofilm was confirmed by scanning electron microscopy (SEM).     

Versatility of merocyanine 540 for the flow cytometric detection of apoptosis in human and murine cells

Since apoptosis plays many roles in development, immune function, and disease, there is an ongoing need to identify inexpensive and reliable fluorochromes for the quantitation of apoptosis. Merocyanine 540 (MC540) binds to the outer membrane of cells and readily fluoresces in the highly disordered membranes of apoptotic cells making them readily detectable by flow cytometry. Protocols for the effective labeling and gating of MC540br apoptotic cells are provided. For example, MC540br cells from dexamethasone (Dex) treated thymocytes were found to be equivalent in proportion to apoptotic cells noted in the propidium iodide (PI) stained and annexin-V stained populations. Sorting of the MC540br cells followed by counterstaining with PI demonstrated that these cells resided in the low DNA fluorescent or sub-G1 region and were small in size based on light scatter. Dexamethasone, etoposide, irradiation, and a calcium ionophore were used to induce cell death with equivalent numbers of apoptotic cells obtained with MC540 and PI. Moreover, apoptotic human bone marrow (BM) B cells, neutrophils, Jurkat T cells, and testicular cells could readily be identified with MC540. The latter is particularly noteworthy since some of the standard methods for identifying cell death have not worked well with human cells. The versatility of this dye is such that it was also possible to phenotypically label cells stained with MC540 to analyze apoptosis in heterogenous populations of cells. Finally, the rate of detection of apoptotic cells after treatment of thymocytes with dexamethasone at 2, 4, 6, and 8 h with MC540 was shown to be equivalent to PI and annexin-V. Taken together, the data demonstrate that when proper precautions are taken, MC540 is a reliable, versatile, and inexpensive fluorochrome that can be used to identify apoptotic cells of human or murine origin even in heterogenous populations that require multicolor labeling.     

BPD-MA光动力作用诱导兔血管平滑肌细胞凋亡的初步研究

目的：探讨新型光敏剂苯并卟啉衍生物单环酸A（BPD-MA）光动力作用诱导血管平滑肌细胞凋亡及机制。方法：用体外培养的兔血管平滑肌细胞，加浓度0．25mg／ml的光敏剂BPD-MA，经能量密度4．8J／cm^2波长650nm半导体激光照射。免疫组化染色鉴定平滑肌细胞，HE染色观察细胞形态，MTT法测定细胞增殖活性，TUNEL法观察细胞凋亡。结果：在4．8J／cm^2激光能量密度照射下，与空白对照组比较，光动力作用对平滑肌细胞增殖活性有显著地抑制作用（P〈0．01），单纯激光组照射埘平滑肌细胞的增殖则无明显抑制作用（P〉0．05）。BPD-MA光动力作用使光动力组的大多数细胞出现凋亡。结论：凋亡可能是苯并卟啉衍生物单环酸A光动力抑制兔血管平滑肌细胞增殖的关键因素。     

High-resolution scanning electron microscopic evaluation of cell-membrane porosity by ultrasound

The effect of low-intensity ultrasound on HL-60 cells (human promyelocytic leukemia cells) in the presence of the photo sensitizing drug merocyanine 540 (MC 540) was evaluated morphologically, using a high-resolution scanning electron microscope and a transmission electron microscope. Exposure of HL-60 cells to ultrasound without MC 540 resulted in a decrease of finger-like processes in the cells. The cells showed many undulating ruffles on the surface. Distinct pits or holes in the membrane were not observed in these cells. The surface of HL-60 cells treated only with MC 540 was relatively smooth compared with that of control cells. HL-60 cells exposed to ultrasound in the presence of MC 540 showed apparent surface deformation. Numerous crater-like depressions of heterogeneous dimensions were observed in many cells. In addition, various-sized pores were noted in the cell membranes of more damaged cells. These results indicate that cell degeneration was induced by a rapid change in cell membrane porosity during sonication in the presence of MC540.     

Stability enhanced polyelectrolyte-coated gold nanorod-photosensitizer complexes for high/low power density photodynamic therapy

Photodynamic therapy (PDT) is a promising treatment modality for cancer and other malignant diseases, however safety and efficacy improvements are required before it reaches its full potential and wider clinical use. Herein, we investigated a highly efficient and safe photodynamic therapy procedure by developing a high/low power density photodynamic therapy mode (high/low PDT mode) using methoxypoly(ethylene glycol) thiol (mPEG-SH) modified gold nanorod (GNR)-AlPcS4 photosensitizer complexes. mPEG-SH conjugated to the surface of simple polyelectrolyte-coated GNRs was verified using Fourier transform infrared spectroscopy; this improved stability, reduced cytotoxicity, and increased the encapsulation and loading efficiency of the nanoparticle dispersions. The GNR-photosensitizer complexes were exposed to the high/low PDT mode (high light dose = 80 mW/cm² for 0.5 min; low light dose = 25 mW/cm² for 1.5 min), and a high PDT efficacy leads to approximately 90% tumor cell killing. Due to synergistic plasmonic photothermal properties of the complexes, the high/low PDT mode demonstrated improved efficacy over using single wavelength continuous laser irradiation. Additionally, no significant loss in viability was observed in cells exposed to free AlPcS4 photosensitizer under the same irradiation conditions. Consequently, free AlPcS4 released from GNRs prior to cellular entry did not contribute to cytotoxicity of normal cells or impose limitations on the use of the high power density laser. This high/low PDT mode may effectively lead to a safer and more efficient photodynamic therapy for superficial tumors.     

Noninvasive optoacoustic temperature determination at the fundus of the eye during laser irradiation

In all fundus laser treatments of the eye, the temperature increase is not exactly known. In order to optimize treatments, an online temperature determination is preferable. We investigated a noninvasive optoacoustic method to monitor the fundus temperature during pulsed laser irradiation. When laser pulses are applied to the fundus, thermoelastic pressure waves are emitted, due to thermal expansion of the heated tissue. Using a constant pulse energy, the amplitude of the pressure wave increases linearly with an increase in the base temperature of between 30 and 80 degrees C. This method was evaluated in vitro on porcine retinal pigment epithelium (RPE) cell samples and clinically during selective RPE treatment with repetitive microsecond laser pulses. During the irradiation of porcine RPE with a neodymium-doped yttrium (Nd:YLF) laser (527 nm, 1.7 micros, 500 Hz repetition rate, 160 mJ/cm(2)) an increase in the base temperature of 30+/-4 degrees C after 100 pulses was found. During patient treatments, a temperature increase of 60+/-11 degrees C after 100 pulses with a 500-Hz repetition rate and 7+/-1 degrees C after 30 pulses with 100 Hz at 520 mJ/cm(2) was found. All measured data were in good agreement with heat diffusion calculations. Optoacoustic methods can be used to noninvasively determine retinal temperatures during pulsed laser treatment of the eye. This technique can also be adapted to continuous-wave photocoagulation, photodynamic therapy and transpupillary thermotherapy, or other fields of laser-heated tissue.     

Pulsed diode laser-based monitor for singlet molecular oxygen

Photodynamic therapy (PDT) is a promising cancer treatment. PDT uses the affinity of photosensitizers to be selectively retained in malignant tumors. When tumors, pretreated with the photosensitizer, are irradiated with visible light, a photochemical reaction occurs and tumor cells are destroyed. Oxygen molecules in the metastable singlet delta state O2(1Delta) are believed to be the species that destroys cancerous cells during PDT. Monitoring singlet oxygen produced by PDT may lead to more precise and effective PDT treatments. Our approach uses a pulsed diode laser-based monitor with optical fibers and a fast data acquisition system to monitor singlet oxygen during PDT. We present results of in vitro singlet oxygen detection in solutions and in a rat prostate cancer cell line as well as PDT mechanism modeling.     

研究不同光敏剂诱导的光动力疗法对人白血病细胞的杀伤作用

目的 研究并比较3种卟啉类光敏剂——血卟啉衍生物(HpD)、癌光啉(PsD007)和血卟啉 单甲醚(HMME)诱导的光动力疗法(PDT)对白血病细胞K562的杀伤效应.方法 以人白血病细胞K562为研究对象,分为对照组和PDT组,以梯度浓度的光敏剂与K562细胞共同孵育,经不同能量光照后,用噻唑蓝(MTT)法测定PDT对K562细胞的杀伤作用.结果 与对照组相比,PDT对K562细胞有明显杀伤作用,并随着光敏剂浓度的增加和光照能量的增大,效果增强.PsD007-PDT和HMME-PDT的效果都明显优于HpD-PDT(P＜0.05);而当光敏剂质量浓度较大(25 μg/ml)或能量密度较大(7.2 J/cm2)时,PsD007-PDT的作用效果优于HMME-PDT.结论 PDT对人白血病细胞K562具有明显的杀伤作用,其对细胞的抑制率具有显著的剂量效应关系;PDT对K562的杀伤效应与光敏剂种类有关,HpD-PDT的杀伤效果不如PsD007和HMME;在较高能量密度和较大光敏剂浓度的条件下,PsD007-PDT的效果优于HMME-PDT.     

Determination of the tumor tissue optical properties during and after photodynamic therapy using inverse Monte Carlo method and double integrating sphere between 350 and 1000 nm

Photodynamic therapy (PDT) efficacy depends on the amount of light distribution within the tissue. However, conventional PDT does not consider the laser irradiation dose during PDT. The optical properties of biological tissues (absorption coefficient μ(a), reduced scattering coefficient μ's), anisotropy factor g, refractive index, etc.) help us to recognize light propagation through the tissue. The goal of this paper is to acquire the knowledge of the light propagation within tissue during and after PDT with the optical property of PDT-performed mouse tumor tissue. The optical properties of mouse tumor tissues were evaluated using a double integrating sphere setup and the algorithm based on the inverse Monte Carlo method in the wavelength range from 350 to 1000 nm. During PDT, the μ(a) and μ's were not changed after 1 and 5 min of irradiation. After PDT, the μ's in the wavelength range from 600 to 1000 nm increased with the passage of time. For seven days after PDT, the μ's increased by 1.7 to 2.0 times, which results in the optical penetration depth decreased by 1.4 to 1.8 times. To ensure an effective procedure, the adjustment of laser parameters for the decreasing penetration depth is recommended for the re-irradiation of PDT.     

Real-time optoacoustic monitoring of vascular damage during photodynamic therapy treatment of tumor

The optoacoustic technique is a noninvasive imaging method with high spatial resolution. It potentially can be used to monitor anatomical and physiological changes. Photodynamic therapy (PDT)-induced vascular damage is one of the important mechanisms of tumor destruction, and real-time monitoring of vascular changes can have therapeutic significance. A unique optoacoustic system is developed for neovascular imaging during tumor phototherapy. In this system, a single-pulse laser beam is used as the light source for both PDT and for concurrently generating ultrasound signals for optoacoustic imaging. To demonstrate its feasibility, this system is used to observe vascular changes during PDT treatment of chicken chorioallantoic membrane (CAM) tumors. The photosensitizer used in this study is protoporphyrin IX (PpIX) and the laser wavelength is 532 nm. Neovascularization in tumor angiogenesis is visualized by a series of optoacoustic images at different stages of tumor growth. Damage of the vascular structures by PDT is imaged before, during, and after treatment. Rapid, real-time determination of the size of targeted tumor blood vessels is achieved, using the time difference of positive and negative ultrasound peaks during the PDT treatment. The vascular effects of different PDT doses are also studied. The experimental results show that a pulsed laser can be conveniently used to hybridize PDT treatment and optoacoustic imaging and that this integrated system is capable of quantitatively monitoring the structural change of blood vessels during PDT. This method could be potentially used to guide PDT and other phototherapies using vascular changes during treatment to optimize treatment protocols, by choosing appropriate types and doses of photosensitizers and doses of light.     

苯并卟啉衍生物单环酸A光动力作用对血管平滑肌细胞增殖的影响

目的：探讨苯并卟啉衍生物单酸环A（BPD—MA）光动力作用对血管平滑肌细胞增殖的影响。方法：用不同浓度的光敏剂BPD-MA作用于体外培养的兔血管平滑肌细胞，经波长650nm半导体激光以能量密度1．2J／cm^2、2．4J／cm^2、4．8J／cm^2。照射，MTT法检测细胞增殖抑制率，用PCNA观察细胞增殖。结果：在1．2J／cm^2．2．4J／cm^2,4．8J／cm^2激光照射下，与空白对照组比较，BPD-MA光动力对平滑肌细胞有显著地抑制作用（P〈0．01），单纯激光组照射对平滑肌细胞的增殖无明显抑制作用（P〉0．05）。PCNA检测显示光动力作用抑制细胞增殖。结论：：BPD—MA光动力作用对血管平滑肌细胞增殖有明显抑制作用。     

Merocyanine 540 as a flow cytometric probe of membrane lipid organization in leukocytes

Merocyanine 540 (MC540) is a fluorescent probe that binds preferentially to membranes with loosely packed lipids. When combined with flow cytometry, it provides a novel methodology for rapidly and quantitatively assessing lipid organization in individual leukocytes. Analysis of cells stained simultaneously with MC540 and 1-[4-trimethylammoniumphenyl]-6-phenyl-1,3,5-hexatriene, to normalize for surface area, revealed that all leukocytes in peripheral blood bind equivalent amounts of dye per unit surface area. This result indicates that the lipids of the plasma membranes of all types of circulating cells are organized similarly. Upon activation with appropriate stimuli, lymphocytes, monocytes, and neutrophils all bound increased amounts of dye per unit surface area, indicating a change in lipid organization to a less-ordered state. Cells stained with MC540 were sorted on the basis of their fluorescence intensity yielding populations homogeneous with respect to lipid organization. Thus not only can MC540 and flow cytometry be combined for analyzing the organization of lipids in individual leukocytes, but sorted populations can be obtained for further biochemical, structural, and functional analyses.     

Photodynamic therapy induces apoptosis in intimal hyperplastic arteries

Photodynamic therapy (PDT) generates free radicals through the absorption of light by photosensitizers. PDT shows promise in the treatment of intimal hyperplasia, which contributes to restenosis, by completely eradicating cells in the vessel wall. This study investigates the mechanisms of PDT-induced cell death. PDT, using the photosensitizer chloroaluminum-sulfonated phthalocyanine (1 mg/kg) and laser light (lambda = 675 nm) 100 J/cm(2) was administered to rat carotid arteries after balloon injury-induced intimal hyperplasia. Apoptosis was determined by cell morphology with light microscopy and transmission electron microscopy, DNA cleavage by terminal dUTP nick-end labeling staining, and nucleosomal fragmentation (ladder pattern) by DNA agarose gel electrophoresis. Four hours after PDT, apoptosis was observed in vascular cells, as evidenced by terminal dUTP nick-end labeling staining and transmission electron microscopy. Within 24 hours no cells were present in the neointima and media. Immunofluorescence using an alpha-smooth muscle cell actin antibody confirmed the disappearance of all neointimal and medial cells within 24 hours. No inflammatory cell infiltrate was observed during this time frame. Apoptosis was sharply confined to the PDT treatment field. These data demonstrate that vascular PDT induces apoptosis as a mechanism of rapid, complete, and precise cell eradication in the artery wall. These findings and the lack of inflammatory reaction provide the basis for understanding and developing PDT for a successful clinical application in the treatment of hyperplastic conditions such as restenosis.     

Induction of apoptosis by hexaminolevulinate-mediated photodynamic therapy in human colon carcinoma cell line 320DM.

Photodynamic therapy (PDT) typically involves systemic or topical administration of a tumor-localizing photosensitizer or prodrug and its subsequent activation by visible light. This results primarily in singlet oxygen-induced photodamage to the tumor. 5-Aminolevulinic acid (ALA) and its derivatives have recently been widely used for PDT due to their selective induction in tumor of endogenous protoporphyrin IX (PpIX), a potent photosensitizer. Although ALA-PDT has achieved successful results in the treatment of several clinical oncological and nononcological diseases, the mechanisms of this modality are still not fully elucidated. In the present study, the human colon carcinoma cell line 320DM was treated in vitro with PDT using hexaminolevulinate (HAL), a hexylester of ALA known to be 50 to 100 times more efficient at producing PpIX formation than ALA itself. PpIX production increased with increasing HAL concentrations in the cells and phototoxicity of the cells was enhanced with increasing light (450 nm) doses. HAL-PDT induced apoptotic cell death, as measured by nuclear staining of Hoechst 33342 for fluorescence microscopy, DNA electrophoresis and TdT staining for flow cytometry. PDT with 5 muM of HAL and a light dose of 640 mJ/cm2 produced a 75% apoptotic cell population 40 hr after the treatment. Furthermore, the loss of mitochondrial membrane potential coincident with the release of cytochrome c from the mitochondria into the cytosol led to a rapid activation of caspase-9 and caspase-3 (an executioner), indicating that the selective damage to the mitochondria by HAL-PDT can induce a cytochrome-c-mediated apoptotic response in the 320DM cells.     

新型卟啉类光敏药物介导的PDT对两种胃癌细胞HGC27和MGC803的杀伤效应

目的 观察本实验室自行合成的新型卟啉类光敏药物对2种人胃癌细胞HGC27和MGC803的光动力学治疗(PDT)作用及作用机制.方法 以人胃癌细胞HGC27和MGC803为实验细胞株.实验分为4组:空白对照组(无药物孵育、无光照),单纯光敏药物组(药物孵育、无光照),单纯光照组(无药物孵育、有光照),光敏药物+光照组(药...     

Selective depletion of a thymocyte subset in vitro with an immunomodulatory photosensitizer

Conventional photodynamic therapy (PDT) utilizes light-absorbing compounds that have anti-cancer activity upon visible light irradiation. PDT has also been utilized for the treatment of certain immune conditions. To further understand the action of PDT upon immune cells, DBA/2 mouse thymocytes were treated with the photosensitizer benzoporphyrin derivative monoacid ring A (BPD-MA, verteporfin) and/or an apoptosis-inducing anti-Fas (APO-1, CD95) monoclonal antibody. Nanomolar levels of BPD-MA in combination with nonthermal visible light irradiation rapidly induced apoptosis as gauged by DNA fragmentation assays. Thymocytes were modestly more sensitive to PDT-induced apoptosis than mature splenic T cells. BPD-MA and light or the anti-Fas antibody decreased CD4(+)CD8(+) cell numbers while relatively sparing CD4(-)CD8(-), CD4(+)CD8(-), and CD4(-)CD8(+) thymocytes. In combination, anti-Fas antibody and PDT augmented activity levels of the apoptosis-related protease caspase-3, cleavage of the caspase-3 substrate poly(ADP) polymerase, and the proportion of cells exhibiting DNA fragmentation and further impacted CD4(+)CD8(+) thymocyte survival. Although CD4(+)CD8(+) thymocytes had the greatest sensitivity to photodynamic depletion, BPD-MA was taken up by the other major thymocyte subsets with equal or greater avidity. Since CD4(+)CD8(+) thymocytes are selectively impacted by PDT and anti-Fas antibody can act in concert with PDT to further cytotoxicity, thymocytes may be useful for the identification of factors that govern immune cell susceptibility to this form of phototherapy.     

HMME介导的光动力疗法诱导鼠肝癌MM45T-Li细胞凋亡的实验研究

目的 探讨光动力疗法对鼠肝癌细胞MM45T-Li凋亡的影响.方法 以血卟啉单甲醚(HMME)为光敏剂,630 nm激光为激发光源的光动力疗法作用于鼠肝癌MM45T-Li细胞.细胞分别与2.5、5、10、20.μg/ml的光敏剂孵育4h后,用不同能量密度的激光照射;MTr法检测HMME的暗毒性以及光动力疗法作用24h后的细胞活性;Hoechst细胞核荧光染色法观察HMME介导的光动力疗法对细胞凋亡的影响.结果 在不照光的情况下各组浓度的HMME对细胞活性无明显抑制作用.当HMME浓度为10、20 μg/ml时,PDT对细胞活性的抑制率随能量密度的增加而增加.Hoechst染色观察PDT作用12h后部分细胞出现染色质凝集、核固缩、核碎裂等形态学改变,5.4 J/cm2及7.2 J/cm2组的凋亡率高于对照组(P＜0.05).结论 HMME介导的光动力疗法可以有效地诱导鼠肝癌MM45T-Li细胞凋亡.     

Hypericin induced death receptor-mediated apoptosis in photoactivated tumor cells

Nasopharyngeal carcinoma (NPC) is a malignant disease of the head/neck region with a 5-year survival level of approximately 65%. To explore the novel therapeutic strategies in the management of this disease, the potential effects of photodynamic therapy (PDT) in NPC cells were investigated. PDT, a new mode of treatment, is based on the combined use of light-absorbing compounds and light irradiation. Two human NPC cells such as, poorly differentiated (NPC/CNE2) and moderately differentiated (NPC/TW0-1) and other types of tumor cells like colon (CCL-220.1) and bladder (SD) undergo rapid apoptosis when treated with PDT sensitized with hypericin (HY). It has been shown that this compound has a strong photodynamic effect on tumors and viruses. However, the initiating events of PDT sensitized HY-induced apoptosis are not identified completely. In this study, we sought to determine whether Fas/FasL upregulation and involvement of mitochondrial events are an early event in HY-treated PDT induced apoptosis. Loss of mitochondrial transmembrane potential, release of cytochrome c, involvement of caspases 8 and 3 and the status caspase-3 specific substrate PARP, were evaluated in PDT treated tumor cells. Photosensitization of HY enhanced both CD95/CD95L expression and induced CD95-signaling dependent cell death in all tumor cell lines studied. CD95/CD95L expression appeared within 2 h following light irradiation and appeared to be a principal event in PDT induced apoptosis. Furthermore, these results indicate that release of mitochondrial cytochrome c into the cytoplasm within 2-3 h post PDT is a secondary event following the activation of initiator caspase-8 preceding Apaf-1, caspase-9 and caspase-3 activation, cleavage of PARP and DNA fragmentation.     

Chlorophyll derivatives (CpD) extracted from silk worm excreta are specifically cytotoxic to tumor cells in vitro

Among chlorophyll derivatives (CpD-A, -B, -C, and -D) extracted from silk worm (Bombyx mori) excretas, CpD-A was extensively studied to clarify its role as a "photosensitizer" for photodynamic therapy (PDT) of tumors in vitro. It was found that CpD-A was photoreactive both in itself and also in its cell bound forms. The cell bound CpD-A produced fluorescent light and singlet oxygen following the exposure to lights of varied wave length. Among them, lights of near 650 nm, which was the maximum absorbance band, efficiently activated CpD-A following the application of only 10 minutes of irradiation. CpD-A was found to have specificity for the human and mouse tumor cells regardless of their species difference. A higher intensity of fluorescence and a larger amount of CpD-A were found in the tumor cells as opposed to the intensity found in normal cells. Only 10 minutes of light irradiation of the CpD-A treated tumor cells resulted in their rapid and complete destruction within 2 hours of irradiation. Simultaneously, more than 80% of the normal human and mouse control cells remained alive after receiving treatment. These findings suggested that CpD-A produced by use of silkworm excreta could be used as a photosensitizer for PDT of tumors by the use of lights of near 650 nm.     

Cell disintegration by laser-induced transient microbubbles and its simultaneous monitoring by interferometry

Selective retina treatment (SRT) is a novel short pulsed laser therapy of several retinal diseases associated with a decreased metabolism at the retinal pigment epithelium (RPE). The range of laser pulse energies is small, in which the desired selective RPE disintegration is achieved without adverse effects to the neural retina. Thus, a real-time dosimetry control is required. We investigated a noninvasive interferometric technique able to monitor microbubble formation around the intracellular melanin granula, which is the origin of the desired RPE damage. A porcine ex vivo RPE model was irradiated by single pulses (350 ns1.7 mus) of a neodymium: yttrium lithium fluoride laser (527 nm). The specimen was simultaneously probed by a Michelson interferometer (helium neon-laser: 633 nm) and by a hydrophone. Cell viability assays (Calcein-AM) were performed after irradiation. At threshold radiant exposure for cell death (ED(50)=129+/-5 mJ cm2 for 350 ns; ED50=180+/-5 mJ cm2 for 1.7 mus), the interferometric transients changed due to microbubble formation. No major differences in the bubble dynamics were observed between both pulse durations. An algorithm to determine cell death from the interferometric transients showed less than 10% false positive or false negative results for the applied laser expositions compared to the viability assay. Interferometry is a reliable noncontact technique to monitor RPE disintegration and may serve as real-time dosimetry control during SRT.