This PDF file contains the front matter associated with SPIE Proceedings Volume 10062, including the Title Page, Copyright information, Table of Contents, and Conference Committee listing.

The mechanism of femtosecond laser nanosurgical attachment is investigated in the following article. Using sub-10 femtosecond laser pulses with 800 nm central wavelength were used to attach retinoblastoma cells. During the attachment process the cell membrane phospholipid bilayers hemifuse into one shared phospholipid bilayer, at the location of attachment. Transmission electron microscopy was used in order to verify the above hypothesis. Based on the imaging results, it was concluded that the two cell membrane coalesce to form one single shared membrane. The technique of cell-cell attachment via femtosecond laser pulses could potentially serve as a platform for precise cell membrane manipulation. Manipulation of the cellular membrane is valuable for studying diseases such as cancer; where the expression level of plasma proteins on the cell membrane is altered.

The current human immunodeficiency virus (HIV) treatment regime possesses the ability to diminish the viral capacity to unnoticeable levels; however complete eradication of the virus cannot be achieved while latent HIV-1 reservoirs go unchallenged. Therapeutic targeting of HIV therefore requires further investigation and current therapies need modification in order to address HIV eradication. This deflects research towards investigating potential novel antiretroviral drug delivery systems. The use of femtosecond (fs) laser pulses in promoting targeted optical drug delivery of antiretroviral drugs (ARVs) into TZMbl cells revolves around using ultrafast laser pulses that have high peak powers, which precisely disrupt the cell plasma membrane in order to allow immediate transportation and expression of exogenous material into the live mammalian cells. A photo-translocation optical setup was built and validated by characterisation of the accurate parameters such as wavelength (800 nm) and pulse duration (115 fs). Optimisation of drug translocation parameters were done by performing trypan blue translocation studies. Cellular responses were determined via cell viability (Adenosine Triphosphate activity) and cell cytotoxicity (Lactate Dehydrogenase) assays which were done to study the influence of the drugs and laser exposure on the cells. After laser irradiation, high cell viability was observed and low toxicity levels were observed after exposure of the cells to both the ARVs and the laser. Our results confirmed that, with minimal damage and high therapeutic levels of ARVs, the fs laser assisted drug delivery system is efficient with benefits of non-invasive and non-toxic treatment to the cells.

Human immunodeficiency virus (HIV-1) infection still remains one amongst the world’s most challenging infections since its discovery. Antiretroviral therapy is the recommended treatment of choice for HIV-1 infection taken by patients orally. The highly active antiretroviral therapy (HAART) prevents the replication of HIV-1 and further destruction of the immune system, therefore enabling the body to fight opportunistic life-threatening infections, cancers, and also arrest HIV infection from advancing to AIDS. The major challenge with HAART is the inability to reach the viral reservoirs where the HIV-1 remains latent and persistent, leading to inability to fully eradicate the virus. This study is aimed at initially designing and assembling a fully functional optical translocation setup to optically deliver antiretroviral drugs into HIV-1 infected cells in a targeted manner using Gaussian beam mode femtosecond laser pulses in-vitro. The main objective of our study is to define the in-vitro drug photo-translocation parameters to allow future design of an efficient drug delivery device with potential in-vivo drug delivery applications. In our experiments, HEK 293T cells were used to produce HIV-1 enveloped pseudovirus (ZM53) to infect TZM-bl cells which were later treated with laser pulses emitted by a titanium sapphire laser (800 nm, 1KHz, 113 fs, ~ 6.5 μW) to create sub-microscopic pores on the cell membrane enabling influx of extracellular media. Following laser treatment, changes in cellular responses were analysed using cell morphology studies, cytotoxicity, and luciferase assay studies. Controls included laser untreated cells incubated with the drug for 72 hours. The data in this study was statistically analysed using the SigmaPlot software version 13.

Cellular manipulation by delivery of molecules into cells has been applied extensively in tissue engineering research for medical applications . The different molecular delivery techniques used range from viral and chemical agents to physical and electrical methods. Although successful in most studies, these techniques have inherent difficulties such as toxicity, unwanted genetic mutations and low reproducibility respectively. Literature recognizes pulsed lasers at femtosecond level to be most efficient in photonic interactions with biological material. As of late, laser pulses have been used for drug and DNA delivery into cells via transient optical perforation of the cellular membrane. Thus in this study, we design and construct an optical system coupled to a femtosecond laser for the purpose of phototransfection or insertion of plasmid DNA (pDNA) into cells using lasers. We used fluorescent green protein (pGFP) to transfect mouse embryonic stem cells as our model. Secondly, we applied fluorescence imaging to view the extent of DNA delivery using this method. We also assessed the biocompatibility of our system by performing molecular assays of the cells post irradiation using adenosine triphosphate (ATP) and lactate dehydrogenase (LDH).

Local resection of early stage tumors in the large bowel via colonoscopy has been a widely accepted surgical modality for colon neoplasm treatment. The conventional electrocautery techniques used for the resection of neoplasia in the mucosal or submucosal layer of colon tissue has been shown to create obvious thermal necrosis to adjacent healthy tissues and lacks accuracy in resection. Ultrafast picosecond (ps) laser ablation using a wavelength of 1030 or 515 nm is a promising surgical tool to overcome the limitations seen with conventional surgical techniques. The purpose of this initial study is to analyze the depth of ablation or the extent of coagulation deployed by the laser as a function of pulse energy and fluence in an ex-vivo porcine model. Precise control of the depth of tissue removal is of paramount importance for bowel surgery where bowel perforation can lead to morbidity or mortality. Thus we investigate the regimes that are optimal for tissue resection and coagulation through plasma mediated ablation of healthy colon tissue. The ablated tissue samples were analyzed by standard histologic methods and a three dimensional optical profilometer technique. We demonstrate that ultrafast laser resection of colonic tissue can minimize the region of collateral thermal damage (<50 μm) with a controlled ablation depth. This surgical modality allows potentially easier removal of early stage lesions and has the capability to provide more control to the surgeon in comparison with a mechanical or electrocautery device.

With continued advancement of solid-state laser technology, high-energy lasers operating in the near-infrared (NIR) band are being applied in an increasing number of manufacturing techniques and medical treatments. Safety-related investigations of potentially harmful laser interaction with skin are commonplace, consisting of establishing the maximum permissible exposure (MPE) thresholds under various conditions, often utilizing the minimally-visible lesion (MVL) metric as an indication of damage. Likewise, characterization of ablation onset and velocity is of interest for therapeutic and surgical use, and concerns exceptionally high irradiance levels. However, skin injury response between these two exposure ranges is not well understood. This study utilized a 1070-nm Yb-doped, diode-pumped fiber laser to explore the response of excised porcine skin tissue to high-energy exposures within the supra-threshold injury region without inducing ablation. Concurrent high-speed videography was employed to assess the effect on the epidermis, with a dichotomous response determination given for three progressive damage event categories: observable permanent distortion on the surface, formation of an epidermal bubble due to bounded intra-cutaneous water vaporization, and rupture of said bubble during laser exposure. ED₅₀ values were calculated for these categories under various pulse configurations and beam diameters, and logistic regression models predicted injury events with approximately 90% accuracy. The distinction of skin response into categories of increasing degrees of damage expands the current understanding of high-energy laser safety while also underlining the unique biophysical effects during induced water phase change in tissue. These observations could prove useful in augmenting biothermomechanical models of laser exposure in the supra-threshold region.

Laser-induced microcavitation refers to the rapid formation and expansion of a vapor bubble inside the bio-tissue when it is exposed to intense, pulsed laser energy. With the associated microscale dissection occurring within the tissue, laserinduced microcavitation is a common approach for high precision bio-surgeries. For example, laser-induced microcavitation is used for laser in-situ keratomileusis (LASIK) to precisely reshape the midstromal corneal tissue through excimer laser beam.

Multiple efforts over the last several years have observed unique characteristics of microcavitions in biotissues. For example, it was found that the threshold energy for microcavitation can be significantly reduced when the size of the biostructure is increased. Also, it was found that the dynamics of microcavitation are significantly affected by the elastic modules of the bio-tissue. However, these efforts have not focused on the early events during microcavitation development.

In this study, a direct numerical simulation of the microcavitation process based on equation of state of the biotissue was established. With the direct numerical simulation, we were able to reproduce the dynamics of microcavitation in water-rich bio tissues. Additionally, an experimental setup in deionized water and 10% PAA gel was made to verify the results of the simulation for early micro-cavitation formation for 10% Polyacrylamide (PAA) gel in deionized water.

A focused laser beam at wavelength of strong water absorption at 1.94 μm can be a good scalpel for precision soft tissue surgery. A fiber Bragg grating-based, all-fiber, continuous-wave as well as modulated, cladding pumped, thulium-doped fiber laser at 1.94 μm has been configured to deliver up to 10 W of laser power under pumping at 793 nm having an efficiency of 32 %. The laser was exposed to freshly sacrificed chicken breast at different power level and exposure time. The formalin-fixed samples were examined by microscopy to identify the ablation region, carbonization and necrosis region for laser parameter optimization.

This study quantifies laser evoked pressure waves in small confined volumes such as a small dish or the cochlea. The pressure was measured with custom fabricated pressure probes in front of the optical fiber. For the pressure measurements during laser stimulation the probes were inserted into scala tympani or vestibuli. At 164 μJ/pulse, the intracochlear pressure was between 96 and 106 dB (re 20 μPa). The pressure was also measured in the ear canal with a sensitive microphone. It was on average 63 dB (re 20 μPa). At radiant energies large enough to evoke an auditory compound action potential, the outer ear canal equivalent pressure was 36-56 dB (re 20 μPa).

Absorption of nanosecond laser pulses induces rapid thermo-elastic deformation in tissue, i.e. a sub-micrometer scale displacement happens within a couple of microseconds. In this study, we initially investigate the depth-resolved deformation using a 1.5 MHz phase-sensitive optical coherence tomography (OCT) system. Functional images can be reconstructed based on the detected deformation, which enables a new imaging modality called thermo-elastic deformation imaging (TDI). Our results show that the associated displacement is related to the optical absorption of the short laser pulses. The TDI images can provide tissue type information in addition to the conventional OCT images.

Short infrared laser pulses induce a variety of effects in cells and tissues, including neural stimulation and inhibition. However, the mechanism behind these physiological effects is poorly understood. It is known that the fast thermal gradient induced by the infrared light is necessary for these biological effects. Therefore, this study tests the hypothesis that the fast thermal gradient induced in a cell by infrared light exposure causes a change in the membrane fluidity. To test this hypothesis, we used the membrane fluidity dye, di-4-ANEPPDHQ, to investigate membrane fluidity changes following infrared light exposure. Di-4-ANEPPDHQ fluorescence was imaged on a wide-field fluorescence imaging system with dual channel emission detection. The dual channel imaging allowed imaging of emitted fluorescence at wavelengths longer and shorter than 647 nm for ratiometric assessment and computation of a membrane generalized polarization (GP) value. Results in CHO cells show increased membrane fluidity with infrared light pulse exposure and this increased fluidity scales with infrared irradiance. Full recovery of pre-infrared exposure membrane fluidity was observed. Altogether, these results demonstrate that infrared light induces a thermal gradient in cells that changes membrane fluidity.

We developed a novel localized antivascular method, namely photo-mediated ultrasound therapy (PUT), by applying synchronized laser and ultrasound pulses. PUT relies on high optical contrast among biological tissues. Taking advantage of the high optical absorption of hemoglobin, PUT can selectively target microvessels without causing unwanted damages to the surrounding tissue. Moreover, PUT working at different optical wavelengths can selectively treat veins or arteries by utilizing the contrast in the optical spectra between deoxy- and oxy-hemoglobin. Through our experiments, we demonstrated that cavitation might have played a key role in PUT. The addition of a laser pulse to an existing ultrasound field can significantly improve the likelihood of inertial cavitation, which can induce microvessel damage through its mechanical effect. In comparison with conventional laser therapies, such as photothermolysis and photocoagulation, the laser energy level needed in PUT is significantly lower. When a nanosecond laser was used, our in vivo experiments showed that the needed laser fluence was in the range of 4 to 40 mJ/cm².

Thermal damage rate processes in biological tissues are usually characterized by a kinetics approach. This stems from experimental data that show how the transformation of a specified biological property of cells or biomolecule (plating efficiency for viability, change in birefringence, tensile strength, etc.) is dependent upon both time and temperature. Here, two disparate approaches were used to study thermal damage rate processes in cultured retinal pigment epithelial cells. Laser exposure (photothermal) parameters included 2-μm laser exposure of non-pigmented cells and 532-nm exposures of cells possessing a variety of melanosome particle densities. Photothermal experiments used a mid-IR camera to record temperature histories with spatial resolution of about 8 μm, while fluorescence microscopy of the cell monolayers identified threshold damage at the boundary between live and dead cells. Photothermal exposure durations ranged from 0.05-20 s, and the effects of varying ambient temperature were investigated. Temperature during heat transfer using a water-jacketed cuvette was recorded with a fast microthermister, while damage and viability of the suspended cells were determined as percentages. Exposure durations for the heat transfer experiments ranged from 50- 60 s. Empirically-determined kinetic parameters for the two heating methods were compared with each other, and with values found in the literature.

To enhance drug delivery performance of drug eluting balloon (DEB) against re-stenosis, we have proposed a heating drug delivery during balloon dilatation using our laser driven short-term thermal angioplasty which may realize to suppress surrounding thermal injury. We studied an influence of vessel dilatation parameters on the heating drug delivery. These parameters were classified into two different forces, that is, circumferential tension and inter-luminal pressure. We think these parameters were not able to determine only by balloon pressure. The circumferential tension with 0—30 mN/mm² was added to a porcine carotid artery using an automatic stage. Various temperature solutions with 37, and 70°C of hydrophobic fluorescent Rhodamine B with 3 μg/ml in concentration were dropped on pig carotid wall. We measured a defined drug delivery amount as well as delivery depth by a microscopic fluorescence measurement on the cross section of the solution delivered vessel. In the case of 37°C, we found the intima surface drug amount with 7 mN/mm² was increased as 10-20 times as other tension cases. On the other hand, at 70°C, we found the optimum tension with 30 mN/mm². We found the drug delivery enhancement might be related to the change of super microscopic surface structure of the vessel. We predict that the collagen thermal denaturation of the vessel wall might play important role to the drug delivery.

To reveal hemolysis phenomena induced by a photosensitization reaction with its environment, we measured absorption spectrum of a blood sample to analyze hemoglobin oxidation and resolved oxygen desorption dynamics. The quartz glass cell with 1 mm optical path length was used as a cuvette. Red blood cell suspension medium of 0.625 hematocrit with 30 μg/ml talaporfin sodium was used as a sample. A red diode laser of 664 nm wavelength was emitted to the cuvette with 120 mW/cm² in irradiance for 40 J/cm². Absorption spectra of the sample were obtained before and after the photosensitization reaction by a spectrophotometer. Multiple regression analysis was employed to obtain concentrations of various hemoglobin species from measured absorption spectrum. Comparing to 0 and 40 J/cm², methemoglobin and deoxygenated hemoglobin concentrations increased 0.19 g/dL and 0.02 g/dL, respectively. Oxygenated hemoglobin concentration decreased 0.17 g/dL. Oxygen environment could also be presented by oxygen pressure calculated from the concentrations of oxygenated hemoglobin and deoxygenated hemoglobin. These obtained hemoglobin concentration changes might indicate hemolysis progress and oxygen environment. We think this simple optical measurement could reveal both the hemolysis and oxygen environment.

Transcranial infrared laser stimulation (TILS) uses infrared light (lasers or LEDs) for nondestructive and non-thermal photobiomodulation on the human brain. Although TILS has shown its beneficial effects to a variety of neurological and psychological conditions, its physiological mechanism remains unknown. Cytochrome-c-oxidase (CCO), the last enzyme in the electron transportation chain, is proposed to be the primary photoacceptor of this infrared laser. In this study, we wish to validate this proposed mechanism. We applied 8 minutes in vivo TILS on the right forehead of 11 human participants with a 1064-nm laser. Broad-band near infrared spectroscopy (bb-NIRS) from 740-900nm was also employed near the TILS site to monitor hemodynamic and metabolic responses during the stimulation and 5-minute recovery period. For rigorous comparison, we also performed similar 8-min bb-NIR measurements under placebo conditions. A multi-linear regression analysis based on the modified Beer-Lambert law was performed to estimate concentration changes of oxy-hemoglobin (Δ[HbO]), deoxy-hemoglobin (Δ[Hb]), and cytochrome-c-oxidase (Δ[CCO]). We found that TILS induced significant increases of [CCO], [HbO] and a decrease of [Hb] with dose-dependent manner as compared with placebo treatments. Furthermore, strong linear relationships or interplays between [CCO] versus [HbO] and [CCO] versus [Hb] induced by TILS were observed in vivo for the first time. These relationships have clearly revealed close coupling/relationship between the hemodynamic oxygen supply and blood volume versus up-regulation of CCO induced by photobiomodulation. Our results demonstrate the tremendous potential of bb-NIRS as a non-invasive in vivo means to study photobiomodulation mechanisms and perform treatment evaluations of TILS.

In order to study cardiomyocyte electrical conduction damage by a photosensitization reaction (PR) mostly comes from outside of the cells in a few minutes after the PR, we studied propagation delay of contact action potential with cardiomyocyte by the PR. To determine appropriate PR condition for tachyarrhythmia ablation, a precise electrophysiological experiment in vitro has been preferable. We measured the contact action potential using a microelectrode array system of which information may be correct than conventional Ca²⁺ measurement. We investigated the propagation delays of an evoked potential to evaluate the electrical conduction damage by the PR. Rat cardiomyocytes were cultivated for 5-7 days on a dish with which 64 electrodes were patterned, in an incubator controlled to 37°C, 5% CO₂. The following conditions were used for the PR: 40 μg/ml talapordfin sodium and 290 mW/cm², 40-78 J/cm² for an irradiation. A 2D map was obtained to visualize the propagation delays of the evoked potential. The propagation speed, which was calculated based on the measured propagation delays, was decreased by about 30-50% on average of all electrodes after the PR. Therefore, we think 2D propagation delays measurement of the evoked potential with contact action potential measuring system might be available to evaluate the acute electrical conduction damage of cardiomyocyte by the PR.

It is reported that the albumin has different structure among animal species. We have proposed a new methodology of cardiac ablation using talaporfin sodium-induced photosensitization reaction with short drug-light interval to realize immediate and permanent therapeutic effect by singlet oxygen production mainly in the interstitial space. The photosensitization reaction efficacy with different animal species should be investing to consider the optimal animal therapeutic model to evaluate the therapeutic effect of new cardiac ablation methodology. We studied the cell-killing efficacy of extracellular talaporfin sodium-induced photosensitization reaction using talaporfin sodium on myocardial cells in vitro with different albumin animal species: human, canine, bovine, and porcine serum albumin. We obtained that the albumin concentration tendency on the binding ratio and cell lethality was different among the animal species but there was no correlation between binding ratio and cell lethality. We found that the cell lethality dependence on albumin concentration showed 2 different groups, human-canine and bovine-porcine. We think that the canine might be useful as a therapeutic animal model since the cytotoxicity tendency on albumin concentration was similar with that of human albumin. These cell lethality tendency difference would be suggested to explain by the existence of the diazepam site that talaporfin sodium binds mainly.

Transcranial infrared laser stimulation (TILS) is a non-destructive and non-thermal photobiomodulation therapy or process on the human brain; TILS uses infrared light from lasers or LEDs and has gained increased recognition for its beneficial effects on a variety of neurological and psychological conditions. While the mechanism of TILS has been assumed to stem from cytochrome-c-oxidase (CCO), which is the last enzyme in the electron transportation chain and is the primary photoacceptor, no literature is found to report electrophysiological response to TILS. In this study, a 64-channel electroencephalography (EEG) system was employed to monitor electrophysiological activities from 15 healthy human participants before, during and after TILS. A placebo experimental protocol was also applied for rigorous comparison. After recording a 3-minute baseline, we applied a 1064-nm laser with a power of 3.5W on the right forehead of each human participant for 8 minutes, followed by a 5-minute recovery period. In 64-channel EEG data analysis, we utilized several methods (root mean square, principal component analysis followed by independent component analysis, permutation conditional mutual information, and time-frequency wavelet analysis) to reveal differences in electrophysiological response to TILS between the stimulated versus placebo group. The analyzed results were further investigated using general linear model and paired t-test to reveal statistically meaningful responses induced by TILS. Moreover, this study will provide spatial mapping of human electrophysiological and possibly neural network responses to TILS for first time, indicating the potential of EEG to be an effective method for monitoring neurological improvement induced by TILS.

Publisher’s Note: This conference presentation, originally published on 4/19/17, was withdrawn per author request.

The immune system is a very complex system that comprises a network of genetic and signaling pathways subtending a network of interacting cells. The location of the cells in a network, along with the gene products they interact with, rules the behavior of the immune system. Therefore, there is a great interest in understanding properly the role of a cell in such networks to increase our knowledge of the immune system response. In order to acquire a better understanding of these processes, cell printing with high spatial resolution emerges as one of the promising approaches to organize cells in two and three-dimensional patterns to enable the study the geometry influence in these interactions. In particular, laser assisted bio-printing techniques using sub-nanosecond laser sources have better characteristics for application in this field, mainly due to its higher spatial resolution, cell viability percentage and process automation. This work presents laser assisted bio-printing of antigen-presenting cells (APCs) in two-dimensional geometries, placing cellular components on a matrix previously generated on demand, permitting to test the molecular interactions between APCs and lymphocytes; as well as the generation of two-dimensional structures designed ad hoc in order to study the mechanisms of mobilization of immune system cells. The use of laser assisted bio-printing, along with APCs and lymphocytes emulate the structure of different niches of the immune system so that we can analyse functional requirement of these interaction.

Corneal biomechanics plays an important role in determining the eye’s structural integrity, optical power and the overall quality of vision. It also plays an increasingly recognized role in corneal transplant and refractive surgery, affecting the predictability, quality and stability of final visual outcome [1]. A critical limitation to increasing our understanding of how corneal biomechanics controls corneal stability and refraction is the lack of non-invasive technologies that microscopically measure local biomechanical properties, such as corneal elasticity within the 3D space. Bubble based acoustic radiation force elastic microscopy (ARFEM) introduce the opportunity to measure the inhomogeneous elastic properties of the cornea by the movement of a micron size cavitation bubble generated by a low energy femtosecond laser pulse [2, 3]. Laser induced breakdown spectroscopy (LIBS) also known as laser induced plasma spectroscopy (LIPS) or laser spark spectrometry (LSS) is an atomic emission spectroscopy [4]. The LIBS principle of operation is quite simple, although the physical processes involved in the laser matter interaction are complex and still not completely understood. In one sentence for description, the laser pulses are focused down to a target so as to generate plasma that vaporizes a small amount of material which the emitted spectrum is measured to analysis the elements of the target.

In the current report we present further developments of a unified Monte Carlo-based computational model and explore hyperspectral modelling of light interaction with volumetrically inhomogeneous scattering tissue-like media. The developed framework utilizes voxelized representation of the medium and considers spatial/volumetric variations in both structural e.g. surface roughness and wavelength-dependant optical properties. We present the detailed description of algorithms for modelling of light-medium interactions and schemes used for voxel-to-voxel photon packet transitions. The results of calculation of diffuse reflectance and Bidirectional Scattering-Surface Reflectance Distribution Function (BSSRDF) are presented. The results of simulations are compared with exact analytical solutions, phantom studies and measurements obtained by a low-cost experimental system developed in house for acquiring shape and subsurface scattering properties of objects by means of projection of temporal sequences of binary patterns. The computational solution is accelerated by the graphics processing units (GPUs) and compatible with most standard graphics/ and computer tomography file formats.

The applications of nanoparticles in optical techniques of diagnosis and treatment of biological tissues are increasing. Image contrast can be improved in diagnostic approaches such as fluorescence, spectroscopy or optical coherence tomography. The therapeutic effect can be increased if nanoparticles are previously incorporated in the biological tissue. This is the case in thermotherapy, or in Photodynamic Therapy. All these applications take advantage of specific properties of the nanoparticles involved, either optical up- or down-conversion, thermal confinement or the ability to act as a drug-carrier.

Although many biomedical applications that involve nanoparticles are being proposed and tested, there is a need to take into account the influence of those nanoparticles on optical radiation propagation. The previously mentioned optical treatment and diagnosis techniques assume a particular optical propagation pattern, which is altered by the addition of nanoparticles. This change depends on the nanoparticle material, shape, size and concentration, among other parameters. In order to try to quantify these changes, in this work several phantoms that include different nanoparticles are analyzed, in order to estimate the influence of nanoparticles in optical propagation. A theoretical model of optical propagation, which takes into account the absorption and scattering changes in the medium, is also considered. Nanoparticles of different sizes from 40 nm to 1 μm are analyzed. Nanoparticle materials of interest in biomedical applications are employed. The results are relevant in diagnosis interpretation of images and treatment outcome evaluation when nanoparticles are present.

The interstitial photodynamic therapy (iPDT) with 5-aminolevulinic acid (5-ALA) is a safe and feasible treatment modality of malignant glioblastoma. In order to cover the tumour volume, the exact position of the light diffusers within the lesion is needed to decide precisely. The aim of this study is the development of evaluation method of treatment volume with 3D Monte Carlo simulation for iPDT using 5-ALA. Monte Carlo simulations of fluence rate were performed using the optical properties of the brain tissue infiltrated by tumor cells and normal tissue. 3-D Monte Carlo simulation was used to calculate the position of the light diffusers within the lesion and light transport. The fluence rate near the diffuser was maximum and decreased exponentially with distance. The simulation can calculate the amount of singlet oxygen generated by PDT. In order to increase the accuracy of simulation results, the parameter for simulation includes the quantum yield of singlet oxygen generation, the accumulated concentration of photosensitizer within tissue, fluence rate, molar extinction coefficient at the wavelength of excitation light. The simulation is useful for evaluation of treatment region of iPDT with 5-ALA.

Both consist of collagen fibrils, sclera is opaque whereas cornea is transparent for optical wavelengths. By employing the pseudospectral time-domain (PSTD) simulation technique, we model light impinging upon cornea and sclera, respectively. To analyze the scattering characteristics of light, the cornea and sclera are modeled by different sizes and arrangements of the non-absorbing collagen fibrils. Various factors are analyzed, including the wavelength of incident light, the thickness of the scattering media, position of the collagen fibrils, size distribution of the fibrils.

The scattering phase function (the probability distribution of the scattering angle) is intimately associated with the cellular organization and ultrastructure of tissue. Since these physical parameters change during e.g. carcinogenesis; quantification of the phase function and related parameters may allow for improved non-invasive, in vivo discrimination between healthy and diseased tissue. Furthermore, for the derivation of models to interpret measured optical signals, assumptions about the phase function of tissue are often made – regularly assuming a Modified Henyey Greenstein. However, in contrast to other optical properties, the phase function has not yet been extensively measured for different tissue types. With conventional goniometers, the exact backscatter direction of 180 degrees cannot be measured. Especially for techniques that detect backscattered light – such as Optical Coherence Tomography and Elastic Scattering spectroscopy – the details of the backward part of the phase function will have a considerable impact on the measured signal. We have therefore developed a setup that can measure the backward part of the phase function: 134 to 180 degrees. Our design is based on full field Optical Coherence Tomography. We detect all angles simultaneously with a camera, while scanning the reference mirror. The phase function scales with the amplitude of the OCT signal for each angle. We will show our results for validation measurements on two silica bead samples of 200 nm and 400 nm beads.

Photodynamic therapy (PDT) is a very effective technique for treatment of certain types of cancer, among the most common, skin cancer. PDT requires the presence of three elements: the photosensitizer, light and oxygen. Penetration depth of light into the tumor depends on both the characteristics of the tissue to be treated and the wavelength. As the light dose to be delivered in each lesion depends on the optical properties of the tissue, all the effects that change these properties should be considered in order to choose suitable doses. There are some studies that have determined the maximum dose of radiation tolerated for certain types of skin, but the influence of the temperature on the optical properties, especially for darker skin types, remains unknown. In this study, we analyzed the optical properties of skin in vivo of different Latin volunteers in order to study the influence of the temperature on the optical properties and thereby to define more precisely the dose of light to be received by each patient in a personalized way. The optical properties of skin in vivo were investigated using an optical system that included an integrating sphere, a tungsten lamp and a spectrophotometer. Such experimental set up-allowed to obtain spectra reflectance of various volunteers and from this measurement, the absorption coefficient was recovered by Inverse Adding Doubling (IAD) program.

The effective scattering Mueller matrices obtained by the simulation were simplified to the reduced matrices and factorized using the Lu-Chipman polar decomposition, which afforded the polarization parameters in two dimensions. In general, the scalar retardance around the illumination point of a pencil beam shows a broad azimuthal dependence with an offset. Photons may behave quite differently under the birefringence according to their polarization state. In contrast, when the birefringence is oriented along the y axis in the plane parallel to the surface (x-y) plane, for example, the azimuthal dependence of the scalar retardance shows clear maxima along the x and y axes and sharp valleys between the maxima. Photons propagating in the medium probably experience the retardance in nearly the same way, when they are polarized linearly and circularly. Moreover, the polarization parameters generally become nonsymmetric with respect to the plane perpendicular to both the x-y plane and the plane containing the birefringence axis, which suggests that the pathway of the lateral propagation of photons from the illumination point to the surrounding is slightly oblique upward relative to the x-y plane.

Skin wrinkles are visually perceived by consumers but they are also known to possess an underlying structure not apparent at the surface of the skin. This underlying structure can be brought out by polarized hyperspectral imaging. Wrinkle patterns of eye crow’s feet are used as example to show a deeper existing pattern and their characterization versus age on a group of volunteers. The skin inhomogeneity changes within each layer of the skin and can be observed in the shorter wavelength region of the spectrum, about 450nm to 500nm which are well suited to image skin surface inhomogeneities within the central and deep epidermis. Imaging in the 550nm range can serve as a larger scale topology reference because of its deeper penetration into the upper dermis. This serves to bring out the underlying wrinkle pattern as imprinted by collagen anisotropies around deep folds but unapparent to the eye yet. The approach has potential applications in evaluating the internal skin patterns non visible to the eye by mapping their spectral dispersion. This method has thus potentials to evaluate the extent of subsurface structures such as acne and other scars and thereby the efficacy of treatments.

A model of interactions between light and skin tissue was reviewed. For the present study the skin of newborns was examined. The characteristics of newborns skin tissue were taken into account when modeling. In the developed model the skin was introduced in three layers: the epidermis, the basal layer and the dermis. The thickness of the skin layers corresponds with the structure of newborns skin. Absorbance of each layer in the visible and near infrared regions of the spectrum was determined by the absorption of three main skin chromophores: blood, melanin and water. The formula of the scattering and absorption coefficients of blood is given in this study. This paper presents the study of the blood oxygenation effect on the signal of diffusely scattered radiation for three distances between the source and the receiver of radiation: 0.3 mm, 0.6 mm and 1.5 mm. The calculation was obtained using the Monte Carlo mathematical modeling. A detailed description of the model is given. The adequacy of the suggested model has been tested by comparing calculated characteristic with the experimental results obtained by means of double integral sphere. The results show that the wavelength range which provides sufficiently accurate measurements depends on the distance between the source and the receiver of radiation and certain data is provided. For the distance of 0.3 mm this range is at 700-780 nm, 950-1000 nm; for 0.6 mm it is at 640-670 nm, 760-780 nm, and 850-870 nm; for 1.5 mm at 620-740 nm.

Laser-induced therapies include laser ablation to remove or cut target tissue by irradiating high-power focused laser beam. These laser treatments are widely used tools for minimally invasive surgery and retinal surgical procedures in clinical settings. In this study, we demonstrate laser tissue interaction images of various sample tissues using high resolution Fourier-domain optical coherence tomography (Fd-OCT). We use a Q-switch diode-pumped Nd:YVO4 nanosecond laser (532nm central wavelength) with a 4W maximum output power at a 20 kHz repetition rate to ablate in vitro and in vivo samples including chicken breast and mouse ear tissues. The Fd-OCT system acquires time-series Bscan images at the same location during the tissue ablation experiments with 532nm laser irradiation. The real-time series of OCT cross-sectional (B-scan) images compare structural changes of 532nm laser ablation using same and different laser output powers. Laser tissue ablation is demonstrated by the width and the depth of the tissue ablation from the B-scan images.

The delivery of genetic material and drugs into mammalian cells using femtosecond (fs) laser pulses is escalating rapidly. This novel light based technique achieved through a precise focusing of a laser beam on the plasma membrane is called photoporation. This technique is attained using ultrashort laser pulses to irradiate plasma membrane of mammalian cells, thus resulting in the accumulation of a vast amount of free electrons. These generated electrons react photochemically with the cell membrane, resulting in the generation of sub-microscopic pores on the cell membrane enabling a variety of extracellular media to diffuse into the cell. This study is aimed at critically analysing the “do’s and don’ts” of designing, assembling, and characterising an optical translocation setup using a femtosecond legend titanium sapphire regenerative amplifier pulsed laser (Gaussian beam, 800 nm, 1 kHz, 113 fs, and an output power of 850 mW). The main objective in our study is to determine optical phototranslocation parameters which are compatible to the plasma membrane and cell viability. Such parameters included beam profiling, testing a range of laser fluencies suitable for photoporation, assessment of the beam quality and laser-cell interaction time. In our study, Chinese Hamster Ovary-K1 (CHO-K1) cells were photoporated in the presence of trypan blue to determine optimal parameters for photoporation experiment. An average power of 4.5 μW, exposure time of 7 ms, with a laser beam spot of ~1.1 μm diameter at the focus worked optimally without any sign of cell stress and cytoplasmic bleeding. Cellular responses post laser treatment were analysed using cell morphology studies.

For a given experimental setting, the measured spatially resolved reflectance rapidly drops with decreasing numerical aperture of the detection scheme. Consequently, for detection schemes with small numerical apertures, the computational time of MC simulations required to obtain adequate signal-to-noise ratio of the spatially resolved reflectance can become very long. We mitigate the issue by virtually increasing the numerical aperture of the detection scheme in MC simulations and devise a criterion for robust estimation of its maximum value. By using the proposed methodology, we show that the acceptance angle of a selected imaging system can be virtually increased from 3 to 11 while preserving a low relative error of the simulated spatially resolved reflectance over a wide range of tissue-like optical properties. As a result, a more than eightfold improvement in the computation time is attained.

Non-subjective, minimally-invasive, and quantifying techniques may support development and evaluation of a fibrosis regression treatment. The build-up of extracellular matrix in liver fibrosis may result on changes of the endogenous fluorescence of tissue. In this work, we evaluate the fluorescence excitation/emission matrix in the UV range for several bulk samples of murine hepatic tissue preserved in different media. Chemical changes on tissue, caused by formaldehyde preservation, alter the endogenous fluorescence spectra. To avoid these drawbacks, phosphate-buffered saline (PBS) or Iscove’s Modified Dulbecco’s Medium were used. PBS buffer showed to be the less harmful and cost-effective preservation medium to study the endogenous fluorescence in fibrotic tissue.

The research of seed sowing qualities demonstrates the considerable influence of laser irradiation on seeds of different species that are essential for the forestry of Russian Federation. For experiment, we used seeds of Spruce fir (Pícea ábies) and Siberian larch (Lárix sibírica). The seeds were exposed to radiation of the following wavelengths: 405 nm 500 mW, 450 nm 3000 mW, 532 nm 550 mW, 640 nm 1000 mW. The results show that laser exposure of seeds has positive impact on growth rate, technical germination ability, root formation, and more over on establishment and root formation while grafting. In experiments is obtained increasing germination by 15% and the germination time to 10%.