This PDF file contains the front matter associated with SPIE Proceedings Volume 9887, including the Title Page, Copyright information, Table of Contents, Introduction (if any), and Conference Committee listing.

We present some of our recent investigations in Dental Medicine using Optical Coherence Tomography (OCT). Time Domain (TD), Spectral Domain (SD), and Swept Source (SS) OCT in-house developed systems are being used, for both ex vivo and in vivo investigations in the oral cavity. We study ex vivo the interface between the tooth and the dental sealant and demonstrate the limitations of the X-rays investigations that are now the gold standard for such procedures. Using OCT, defects in the interface that cannot be identified in radiographs can be determined both as position and magnitude. The drilling process of teeth can also be characterized in real time using OCT, to monitor the remaining dentin thickness (RDT) in order to avoid opening the pulp chamber. We demonstrate in this respect that an RDT of 0.5 mm is the minimum value to assure the integrity of the dentin wall between the drilled cavity and the pulp chamber; at an RDT of 0.3 mm or less a fracture is initiated, the dentin is punctured and endodontic treatment must follow. In vivo OCT investigations in the oral cavity were also performed (i.e., for metalloceramic prostheses and for ceramic inlay tooth interfaces), with the low cost, light weight and versatile handheld probes with 1D galvoscanners that we have developed and applied for a range of in-house developed OCT systems, in various clinical applications. They are briefly discussed, as well as some of our current and future work in the field, including for studies of soft tissue in the mouth.

Multimodal optical coherence tomography (OCT) is an emerging tool for tissue state characterization. Optical coherence elastography (OCE) is an approach to mapping mechanical properties of tissue based on OCT. One of challenging problems in OCE is elimination of the influence of residual local tissue relaxation that complicates obtaining information on elastic properties of the tissue. Alternatively, parameters of local relaxation itself can be used as an additional informative characteristic for distinguishing the tissue in normal and pathological states over the OCT image area. Here we briefly present an OCT-based approach to evaluation of local relaxation processes in the tissue bulk after sudden unloading of its initial pre-compression. For extracting the local relaxation rate we evaluate temporal dependence of local strains that are mapped using our recently developed hybrid phase resolved/displacement-tracking (HPRDT) approach. This approach allows one to subtract the contribution of global displacements of scatterers in OCT scans and separate the temporal evolution of local strains. Using a sample excised from of a coronary arteria, we demonstrate that the observed relaxation of local strains can be reasonably fitted by an exponential law, which opens the possibility to characterize the tissue by a single relaxation time. The estimated local relaxation times are assumed to be related to local biologically-relevant processes inside the tissue, such as diffusion, leaking/draining of the fluids, local folding/unfolding of the fibers, etc. In general, studies of evolution of such features can provide new metrics for biologically-relevant changes in tissue, e.g., in the problems of treatment monitoring.

A frequent quantification of drugs concentrations in plasma of patients subject to chemotherapy is seldom performed, mostly because the standard methods (Gas or Liquid Chromatography coupled with Mass Spectroscopy) are expensive and time consuming. In this paper we report the approach pursued in one of the research units of the EU project RAMAN4CLINICS to tackle the problem of a low cost, time adherent quantification of drugs used for oncological patients using a Surface Enhanced Raman Scattering (SERS) spectroscopy. More specifically, the issues concerning the repeatability of the nanostructured substrates will be presented and some promising results to increase the selectivity of the measures toward specific drugs will be discussed, with examples concerning one cytotoxic agent, Irinotecan and one kinase inhibitor, Sunitinib.

We report three strategies of surface enhanced Raman spectroscopy (SERS) for β-hematin and hemozoin detection in malaria infected human blood, which can be potentially developed for field malaria diagnosis. In the first strategy, we used silver coated magnetic nanoparticles (Fe₃O₄@Ag) in combination with an external magnetic field to enhance the Raman signal of β-hematin. Then we developed two SERS methods without the requirement of magnetic field for malaria infection diagnosis. In Method 1, silver nanoparticles were synthesized separately and then mixed with lysed blood just like in traditional SERS measurements; while in Method 2, we developed an ultrasensitive SERS method by synthesizing silver nanoparticles directly inside the parasites of Plasmodium falciparum. Method 2 can be also used to detect single parasites in the ring stage.

Despite several advances in molecular biology and diagnostics, Polymerase Chain Reaction (PCR) is currently the gold standard for nucleic acids amplification and detection, due to its versatility, low-cost and universality, with estimated <10 billion reactions per year and a worldwide market of several billion dollars/year. Nevertheless, PCR still relies on the laborious, time-consuming, and multi-step gel electrophoresis-based detection, which includes gel casting, electrophoretic run, gel staining, and gel visualization. In this work, we propose a “PCR developer”, namely a universal one-step, one-tube method, based on controlled aggregation of gold nanoparticles (AuNPs), to detect PCR products by naked eye in few minutes, with no need for any instrumentation. We demonstrated the specificity and sensitivity of the PCR developer on different model targets, suitable for a qualitative detection in real-world diagnostics (i.e., gene rearrangements, genetically modified organisms, and pathogens). The PCR developer proved to be highly specific and ultra-sensitive, discriminating down to few copies of HIV viral DNA, diluted in an excess of interfering human genomic DNA, which is a clinically relevant viral load. Hence, it could be a valuable tool for both academic research and clinical applications.

Nucleases are ubiquitous enzymes that degrade DNA or RNA, thus they can prejudice the good outcome of molecular biology experiments involving nucleic acids. We propose a colorimetric test for the naked-eye detection of nuclease contaminations. The system uses an hybrid nanosensor, based on gold nanoparticles functionalized with DNA probes. Our assay is rapid, instrument-free, simple and low-cost. Moreover, it reaches sensitivity equal or better than those of commercial kits, and presents a lot of advantageous aspects. Therefore, it is very competitive, with a real market potential. This test will be relevant in routine process monitoring in scientific laboratories, and in quality control in clinical laboratories and industrial processes, allowing the simultaneous detection of nucleases with different substrate specificities and large-scale screening.

We report a simple, rapid and low-cost test, based on gold nanoparticles, for the naked-eye colorimetric detection of a signature of single nucleotide polymorphisms (SNPs) relevant for the personalized medicine of psoriasis patients. We validated the colorimetric assay on real-world DNA samples from a cohort of 30 psoriasis patients and we compared the results, in double-blind, with those obtained with two state-of-the-art instrumental techniques, namely reverse dot blotting and direct sequencing, finding 100% agreement. We demonstrated high accuracy, sensitivity and specificity of the colorimetric test that can be easily adapted for the genotypization of different SNPs, important for the pharmacogenomics of various diseases, and in other fields, such as food traceability and population structure analysis.

The problem of cancer control requires design of new approaches for instrumental diagnostics, as the accuracy of cancer detection on the first step of diagnostics in clinics is slightly more than 50%. In this study, we present a method of visualization and diagnostics of skin and lung tumours based on registration and processing of tissues hyperspectral images. In a series of experiments registration of hyperspectral images of skin and lung tissue samples is carried out. Melanoma, basal cell carcinoma, nevi and benign tumours are studied in skin ex vivo and in vivo experiments; adenocarcinomas and squamous cell carcinomas are studied in ex vivo lung experiments. In a series of experiments the typical features of diffuse reflection spectra for pathological and normal tissues were found. Changes in tissues morphology during the tumour growth lead to the changes of blood and pigments concentration, such as melanin in skin. That is why tumours and normal tissues maybe differentiated with information about spectral response in 500-600 nm and 600 – 670 nm areas. Thus, hyperspectral imaging in the visible region may be a useful tool for cancer detection as it helps to estimate spectral properties of tissues and determine malignant regions for precise resection of tumours.

We propose a compact, self-aligned, low-cost, and versatile infrared diffuse-reflectance laser imaging system using a laser feedback interferometry technique with possible applications in in vivo biological tissue imaging and skin cancer detection. We examine the proposed technique experimentally using a three-layer agar skin phantom. A cylindrical region with a scattering rate lower than that of the surrounding normal tissue was used as a model for a non-melanoma skin tumour. The same structure was implemented in a Monte Carlo computational model. The experimental results agree well with the Monte Carlo simulations validating the theoretical basis of the technique. Results prove the applicability of the proposed technique for biological tissue imaging, with the capability of depth sectioning and a penetration depth of well over 1.2 mm into the skin phantom.

A probabilistic robust diagnostic algorithm is very much essential for successful cancer diagnosis by optical spectroscopy. We report here support vector machine (SVM) classification to better discriminate the colon and cervical cancer tissues from normal tissues based on elastic scattering spectroscopy. The efficacy of SVM based classification with different kernel has been tested on multifractal parameters like Hurst exponent, singularity spectrum width in order to classify the cancer tissues.

Investigation of malignant skin tumors diagnosis was performed involving two setups for native tissues fluorescence control in visible and near infrared regions. Combined fluorescence analysis for skin malignant melanomas and basal cell carcinomas was performed. Autofluorescence spectra of normal skin and oncological pathologies stimulated by 457 nm and 785 nm lasers were registered for 74 skin tissue samples. Spectra of 10 melanomas and 27 basal cell carcinomas were registered ex vivo. Skin tumors analysis was made on the basis of autofluorescence spectra intensity and curvature for analysis of porphyrins, lipo-pigments, flavins and melanin. Separation of melanomas and basal cell carcinomas was performed on the basis of discriminant analysis. Overall accuracy of basal cell carcinomas and malignant melanomas separation in current study reached 86.5% with 70% sensitivity and 92.6% specificity.

Non-contact imaging photoplethysmography (iPPG) to detect pulsatile blood microcirculation in tissue has been selected as a successor to low spatial resolution and slow scanning blood perfusion techniques currently employed by clinicians. The proposed iPPG system employs a novel illumination source constructed of multiple high power LEDs with narrow spectral emission, which are temporally modulated and synchronised with a high performance sCMOS sensor. To ensure spectrum stability and prevent thermal wavelength drift due to junction temperature variations, each LED features a custom-designed thermal management system to effectively dissipate generated heat and auto-adjust current flow. The use of a multi-wavelength approach has resulted in simultaneous microvascular perfusion monitoring at various tissue depths, which is an added benefit for specific clinical applications. A synchronous detection algorithm to extract weak photoplethysmographic pulse-waveforms demonstrated robustness and high efficiency when applied to even small regions of 5 mm². The experimental results showed evidences that the proposed system could achieve noticeable accuracy in blood perfusion monitoring by creating complex amplitude and phase maps for the tissue under examination.

The capacity of pathogens to acquire nutrients from their host cells is one of the most fundamental aspects of infection biology. Hence, measuring the patterns of nutrients’ uptake by pathogens is essential for understanding the interactions of pathogens with eukaryotic host cells. In this study, we optimized a technique that allows fast and non-destructive measurement of the amino acid Phenylalanine (Phe) acquired by the trophozoite stage of the protozoan Acanthamoeba castellanii (A. castellanii) as they engage with individual human retinal pigment epithelial cells (ARPE-19). ARPE-19 host cells were pre-saturated with Deuterated Phe (L-Phe(D8)) to replace the native substrate Phe (L-Phe). The uptake of L-Phe(D8) by A. castellanii trophozoites was measured by Raman microspectroscopy. This approach allowed us to characterize the uptake patterns of this essential amino acid into A. castellanii trophozoites at a single cell level. At 24 hours post infection (PI) A. castellanii trophozoites are capable of salvaging L-Phe(D8) from host cells. The uptake pattern was time-dependent during the first 24 hours of infection and complete substitution with L-Phe(D8) in all parasites was detected at 48 hours PI. On the other hand, isolated A. castellanii trachyzoites (grown without host cells) did not show significant uptake for L-Phe(D8) from the media; only achieved an uptake ratio of 16-18% of L-Phe(D8) from the culture medium after 24 hours. These findings demonstrate the potential of combining Raman microspectroscopy and stable isotope labelling approaches to elucidate the role of metabolism in mediating A. castellanii interaction with host cells.

Here we report on the preparation of uniform Au@Ag core/shell nanorods with a controllable Ag shell thickness. 4- aminothiophenol molecules, used as the Raman reporters, were placed between the Au core and the Ag shell. The dependence of Raman intensity on the inside/surface location of the reporter molecules was studied. The interior molecules showed a strong and uniform Raman intensity, at least an order of magnitude higher than that of the molecules on the nanoparticle surface. In contrast to usual surface-functionalized Raman tags, aggregation and clustering of nanoparticles with embedded molecules decreased the SERS signal. The findings from this study provide the basis for a novel technique of low analyte concentration detection based on high SERS response inside the core/shell metal nanostructures.

Confocal Raman microspectroscopy is a relevant and useful tool to perform in vivo diagnosis of cutaneous tissues noninvasively and without labeling. This optical technique provides in-depth molecular and conformational characterization of skin. Unfortunately, spectral distortions occur due to elastic scattering. Our objective is to correct the attenuation of in-depth Raman peaks intensity by considering elastic scattering in biological tissues. In this purpose, a correction model was constructed using skin scattering properties as parameters thus enabling quantitative analysis. The work presented here is a technique of in vivo Diffuse Reflectance Micro-Spectroscopy called Micro-DRS. It achieves optical properties characterization in the skin layers probed by Raman microspectroscopy. The Micro-DRS setup can easily be coupled to a confocal Raman micro-probe to perform simultaneous measurements. Thanks to Monte Carlo simulations and experimental results obtained on homemade solid phantoms mimicking skin optical properties, we show that it is possible to measure the absorption coefficient μ_a, the reduced scattering coefficient μ_s', the scattering coefficient μ_s and the anisotropy of scattering g with this new apparatus. The measured scattering properties can be used subsequently as parameters in our correction model. Coupled to a Raman micro-spectrometer, Micro-DRS enables a quantitative analysis when tracking drug penetration through skin and it can be used independently to provide additional diagnosing criterions.

Bone diseases and disorders are a growing challenge in aging populations; so effective diagnostic and therapeutic solutions are now essential to manage the demands of healthcare sectors effectively. Spatially offset Raman spectroscopy (SORS) allows for chemically specific sub-surface probing and has a great potential to become an in vivo tool for early non-invasive detection of bone conditions. Bone is a complex hierarchical material and the volume probed by SORS is dependent on its optical properties. Understanding and taking into account the variations in diffuse scattering properties of light in various bone types is essential for the effective development and optimization of SORS as a diagnostic in vivo tool for characterizing bone disease. This study presents SORS investigations at 830 nm excitation on two specific types of bone with differing mineralization levels. Thin slices of bone from horse metacarpal cortex (0.6 mm thick) and whale bulla (1.0 mm thick) were cut and stacked on top of each other (4-7 layers with a total thickness of 4.1 mm). To investigate the depth origin of the detected Raman signal inside the bone a 0.38 mm thin Teflon slice was used as test sample and inserted in between the layers of stacked bone slices. For both types of bone it could be demonstrated that chemically specific Raman signatures different from those of normal bone can be retrieved through 3.8-4.0 mm of overlying bone material with a spatial offset of 7-8 mm. The determined penetration depths can be correlated with the mechanical and optical properties of the specimens. The findings of this study increase our understanding of SORS analysis of bone and thus have impact for medical diagnostic applications e.g. enabling the non-invasive detection of spectral changes caused by degeneration, infection or cancer deep inside the bone matrix.

Dynamics of temperature signal in operation area and laser power at nevus, papilloma, and keratoma in vivo removal by a 980±10 nm diode laser with "blackened" tip operating in continuous (CW) mode and with temperature feedback (APC) mode are presented. Feedback allows maintaining temperature in the area of laser treatment at a preset level by regulating power of diode laser radiation (automatic power control). Temperature in the area of laser treatment was controlled by measuring the amplitude of thermal radiation, which occurs when tissue is heated by laser radiation. Removal of neoplasm was carried out in CW mode with laser radiation average power of 12.5±0.5 W; mean temperature in the area of laser treatment was 900±10°C for nevus, 800±15°C for papilloma, and 850±20°C for keratoma. The same laser radiation maximal power (12.5 W) and targeted temperature (900°C) were set for nevus removal in APC mode. The results of investigation are real time oscillograms of the laser power and temperature in the area of laser treatment at neoplasms removal in two described above modes. Simultaneously with the measurement of laser power and the temperature in the area of laser treatment video recording of surgeon manipulations was carried out. We discuss the correlation between the power of the laser radiation, the temperature in the area of laser treatment and consistency of surgeon manipulation. It is shown that the method of removal (excision with or without traction, scanning) influences the temperature in the area of laser treatment. It was found, that at removal of nevus with temperature feedback (APC) mode to achieve comparable with CW mode temperature in the area of laser treatment (900±10°C) 20-50% less laser power is required. Consequently, removing these neoplasms in temperature feedback mode can be less traumatic than the removal in CW mode.

Phase-resolved measurements found numerous applications in low-coherence methods, in particular in OCT-based compressional elastography, where phase-variation gradients are used for estimating strains produced by the OCT probe pressed onto the tissue. Conventionally, for the reference and deformed pixelated OCT scans, one performs comparison of phases taken from pixels with the same coordinates. This is reasonable in regions of sufficiently small sub-pixel displacements, for which the so-compared pixels contain the same scatterers. Furthermore, to avoid error-prone multiple phase unwrapping for reconstructing displacements, one have to ensure even smaller sub-wavelength displacements. This limits the allowable strains to less than ~10^-4-10^-3, although such weak phase gradients can be strongly corrupted by measurement noises. Here, we discuss how creation of an order of magnitude greater strains can be used for increasing the signal-to noise ratio in estimating phase gradients by obviating the phase-unwrapping procedures and reducing the influence of decorrelation noise for supra-pixel displacements. This optimized phase-variation measurement makes it possible to perform strain mapping in optical coherence elastography with exceptionally high tolerance to noises due to possibility of using significantly increased strains. We also discuss the effect of “frozen-phase zones” associated with displaced strong scatterers. This effect can result in appearance of artifacts in the form of false stiff inclusions in elastograms in the vicinity of bright scatterers in OCT scans. We present analytical arguments, numerical simulations and experimental examples illustrating the above-mentioned features of the “frozen-phase” effect and advantages of using the proposed optimized phase-variation measurement with pixel-scale displacement compensation in the compared OCT scans.

Coronary artery disease (CAD) contributes to millions of deaths each year. The identification of vulnerable plaques is essential to the diagnosis of CAD but is challenging. Molecular probes can improve the detection of these plaques using intravascular imaging methods. Fluorescence lifetime sensing is a safe and robust method to image these molecular probes. We present two variations of an optical system for intravascular near-infrared (NIR) fluorescence lifetime sensing through a multimode fiber. Both systems are built around a recently developed fast and efficient CMOS detector, the current-assisted photonic sampler (CAPS) that is optimized for sub-nanosecond NIR fluorescence lifetime sensing. One system mimics the optical setup of an epifluorescence microscope while the other uses a practical fiber optic coupler to separate fluorescence excitation and emission. We test both systems by measuring the lifetime of several NIR dyes in DMSO solutions and we show that these systems are capable of detecting lifetimes of solutions with concentrations down to 370 nM and this with short acquisition times. These results are compared with time-correlated single photon counting (TCSPC) measurements for reference.

Recently, there are great deals of skin fluorescence studies for diagnostic purposes in medicine. Measurement of the intensity of autofluorescence (AF) is suitable method for diagnostic, because it does not require traumatic procedures. Skin AF is widely used by doctors in order to assess the concentration of advanced glycation endproduct (AGE). There are no in vivo fluorescence meters made in Russia, which are affordable, portable, easy-to-use and easily replicable. This paper is devoted to study of the fluorimeter and its mathematical model of spectral characteristics that were developed by authors. Fluorimeter and its software are fully operational and they were given to doctors for testing in the real clinic conditions in order to get a set of AF statistics for patients.

Fluorescence lifetime imaging microscopy (FLIM) has allowed scientists to discern information about the chemical properties of biological processes and has become a vital tool in the life sciences and medical research communities. Measuring the spatial lifetime distribution of the fluorophores as well as the intensity distribution enables users to discern vital information about the chemical environment. It however, remains challenging and often involves slow scanning. We present a new microscope system based on light sheet illumination that uses a micro channel plate (MCP) device called a Capacitive Division Imaging Readout (CDIR) which has been developed by Photek Ltd. The device uses an array of capacitors to move the charge site from the MCP to four pre-amplifiers and time-over-threshold discriminators. This camera has the ability to image photons as well as measure the arrival time, enabling high speed FLIM imaging of biological samples.

The original UV biodosimeter for an in situ monitoring of the vitamin-D-synthetic capacity of sunlight and/or artificial UV sources is based on the same photoreaction in vitro by which vitamin D is synthesized in human skin from initial provitamin D via photo- and thermo-induced monomolecular isomerizations. Therefore, targets for UV photons in the biodosimeter are the provitamin D molecules embedded in specially designed UV transparent and stable matrix. The dosimeter response to UV radiation is photoinduced conversion of provitamin D into previtamin D which is immediate precursor of vitamin D. Thus, biological ‘antirachitic’ UV dose is determined by the amount of accumulated previtamin D. To follow the photoreaction course in real time three operation modes of varying complexity have been developed.

A new device incorporating a new algorithm and measurement process allows simultaneous noninvasive in vivo monitoring of intravascular plasma volume and red blood cell volume. The purely optical technique involves probing fingertip skin with near infrared laser light and collecting the wavelength shifted light, that is, the inelastic emission (IE) which includes the unresolved Raman and fluorescence, and the un-shifted emission, that is, the elastic emission (EE) which includes both the Rayleigh and Mie scattered light. Our excitation and detection geometry is designed so that from these two simultaneous measurements we can calculate two parameters within the single scattering regime using radiation transfer theory, the intravascular plasma volume fraction and the red blood cell volume fraction. Previously calibrated against a gold standard FDA approved device, 2 hour monitoring sessions on three separate occasions over a three week span for a specific, motionless, and mostly sleeping individual produced 3 records containing a total of 5706 paired measurements of hematocrit and plasma volume. The average over the three runs, relative to the initial plasma volume taken as 100%, of the plasma volume±1σ was 97.56±0.55 or 0.56%.For the same three runs, the average relative hematocrit (Hct), referenced to an assumed initial value of 28.35 was 29.37±0.12 or stable to ±0.4%.We observe local deterministic circulation effects apparently associated with the pressure applied by the finger probe as well as longer timescale behavior due to normal ebb and flow of internal fluids due to posture changes and tilt table induced gravity gradients.

Timely microbiological results are essential to allow clinicians to optimize the prescribed treatment, ideally at the initial stage of the therapeutic process. Several approaches have been proposed to solve this issue and to provide the microbiological result in a few hours directly from the sample such as molecular biology. However fast and sensitive those methods are not based on single phenotypic information which presents several drawbacks and limitations. Optical methods have the advantage to allow single-cell sensitivity and to probe the phenotype of measured cells. Here we present a process and a prototype that allow automated single-bacteria phenotypic analysis. This prototype is based on the use of Digital In-line Holography techniques combined with a specially designed Raman spectrometer using a dedicated device to capture bacteria. The localization of single-cell is finely determined by using holograms and a proper propagation kernel. Holographic images are also used to analyze bacteria in the sample to sort potential pathogens from flora dwelling species or other biological particles. This accurate localization enables the use of a small confocal volume adapted to the measurement of single-cell. Along with the confocal volume adaptation, we also have modified every components of the spectrometer to optimize single-bacteria Raman measurements. This optimization allowed us to acquire informative single-cell spectra using an integration time of 0.5s only. Identification results obtained with this prototype are presented based on a 65144 Raman spectra database acquired automatically on 48 bacteria strains belonging to 8 species.

This work is devoted to study the possibility of plasma proteins (albumin, globulins) concentration measurement using Raman spectroscopy setup. The blood plasma and whole blood were studied in this research. The obtained Raman spectra showed significant variation of intensities of certain spectral bands 940, 1005, 1330, 1450 and 1650 cm^-1 for different protein fractions. Partial least squares regression analysis was used for determination of correlation coefficients. We have shown that the proposed method represents the structure and biochemical composition of major blood proteins.

Food authentication and the detection of adulterated products are recent major issues in the food industry as these topics are of global importance for quality control and food safety. To effectively address this challenge requires fast, reliable and non-destructive analytical techniques. Shifted Excitation Raman Difference Spectroscopy (SERDS) is well suited for identification purposes as it combines the chemically specific information obtained by Raman spectroscopy with the ability for efficient fluorescence rejection. The two slightly shifted excitation wavelengths necessary for SERDS are realized by specially designed microsystem diode lasers. At 671 nm the laser (optical power: 50 mW, spectral shift: 0.7 nm) is based on an external cavity configuration whereas an emission at 783 nm (optical power: 110 mW, spectral shift: 0.5 nm) is achieved by a distributed feedback laser. To investigate the feasibility of SERDS for rapid and nondestructive authentication purposes four types of cheese and three different cheese analogues were selected. Each sample was probed at 8 different positions using integration times of 3-10 seconds and 10 spectra were recorded at each spot. Principal components analysis was applied to the SERDS spectra revealing variations in fat and protein signals as primary distinction criterion between cheese and cheese analogues for both excitation wavelengths. Furthermore, to some extent, minor compositional differences could be identified to discriminate between individual species of cheese and cheese analogues. These findings highlight the potential of SERDS for rapid food authentication potentially paving the way for future applications of portable SERDS systems for non-invasive in situ analysis.

Gold nanocages (AuNCs) have been shown to be a useful tool both for imaging and hyperthermia therapy of cancer, thanks to their outstanding optical properties, low toxicity and facile functionalization with targeting molecules, including peptides and antibodies. In particular, hyperthermia is a minimally invasive therapy which takes advantage of the peculiar properties of gold nanoparticles to efficiently convert the absorbed light into heat. Here, we use AuNCs for the selective targeting and imaging of prostate cancer cells. Moreover, we report the hyperthermic effect characterization of the AuNCs both in solution and internalized in cells. Prostate cancer cells were irradiated at different exposure times, with a pulsed near infrared laser, and the cellular viability was evaluated by confocal microscopy.

Darkfield and confocal laser scanning microscopy both allow for a simultaneous observation of live cells and single nanoparticles. Accordingly, a characterization of nanoparticle uptake and intracellular mobility appears possible within living cells. Single particle tracking makes it possible to characterize the particle and the surrounding cell. In case of free diffusion, the mean squared displacement for each trajectory of a nanoparticle can be measured which allows computing the corresponding diffusion coefficient and, if desired, converting it into the hydrodynamic diameter using the Stokes-Einstein equation and the viscosity of the fluid. However, within the more complex system of a cell’s cytoplasm unrestrained diffusion is scarce and several other types of movements may occur. Thus, confined or anomalous diffusion (e.g. diffusion in porous media), active transport, and combinations thereof were described by several authors. To distinguish between these types of particle movement we developed an appropriate classification method, and simulated three types of particle motion in a 2D plane using a Monte Carlo approach: (1) normal diffusion, using random direction and step-length, (2) subdiffusion, using confinements like a reflective boundary with defined radius or reflective objects in the closer vicinity, and (3) superdiffusion, using a directed flow added to the normal diffusion. To simulate subdiffusion we devised a new method based on tracks of different length combined with equally probable obstacle interaction. Next we estimated the fractal dimension, elongation and the ratio of long-time / short-time diffusion coefficients. These features were used to train a random forests classification algorithm. The accuracy for simulated trajectories with 180 steps was 97% (95%-CI: 0.9481-0.9884). The balanced accuracy was 94%, 99% and 98% for normal-, sub- and superdiffusion, respectively. Nanoparticle tracking analysis was used with 100 nm polystyrene particles to get trajectories for normal diffusion. As a next step we identified diffusion types of nanoparticles in vital cells and incubated V79 fibroblasts with 50 nm gold nanoparticles, which appeared as intensely bright objects due to their surface plasmon resonance. The movement of particles in both the extracellular and intracellular space was observed by dark field and confocal laser scanning microscopy. After reducing background noise from the video it became possible to identify individual particle spots by a maximum detection algorithm and trace them using the robust single-particle tracking algorithm proposed by Jaqaman, which is able to handle motion heterogeneity and particle disappearance. The particle trajectories inside cells indicated active transport (superdiffusion) as well as subdiffusion. Eventually, the random forest classification algorithm, after being trained by the above simulations, successfully classified the trajectories observed in live cells.

The first results concerning the possibility to use Delayed Luminescence spectroscopy to evaluate the in vitro induction of cytotoxic effects on human glioblastoma cells of nanostructured lipid carrier and drug-loaded nanostructured lipid carrier are showed in this contribution. We tested the effects of nanostructured lipid carrier, ferulic acid and ferulic acidloaded nanostructured lipid carrier on U-87MG cell line. The study seems to confirm the ability of Delayed Luminescence to be sensible indicator of alterations induced on functionality of the mitochondrial respiratory chain complex I in U-87MG cancer cells when treated with nanostructured lipid carriers.

A new setup was recently proposed to perform Mueller matrix polarimetry at 100 kHz using a swept laser source, high order retarders and a single channel photodetector. In this communication, we present the implementation of this setup on a laser scanning microscope to perform high speed scanning Mueller microscopy in transmission. Calibration of the instrument is briefly described and precision and stability over time are evaluated. Finally, Mueller images of a manufactured scene are reported. To our best knowledge, this is the first time that Mueller polarimetry is performed using a laser scanning microscope. We further plan to develop confocal/nonlinear/Mueller microscopy from the same setup in order to produce multimodal contrast images of biological samples.

3-D bioconstructions were created using the evaporation method of the water-albumin solution with carbon nanotubes (CNTs) by the continuous and pulsed femtosecond laser radiation. It is determined that the volume structure of the samples created by the femtosecond radiation has more cavities than the one created by the continuous radiation. The average diameter for multi-walled carbon nanotubes (MWCNTs) samples was almost two times higher (35-40 nm) than for single-walled carbon nanotubes (SWCNTs) samples (20-30 nm). The most homogenous 3-D bioconstruction was formed from MWCNTs by the continuous laser radiation. The hardness of such samples totaled up to 370 MPa at the nanoscale. High strength properties and the resistance of the 3-D bioconstructions produced by the laser irradiation depend on the volume nanotubes scaffold forming inside them. The scaffold was formed by the electric field of the directed laser irradiation. The covalent bond energy between the nanotube carbon molecule and the oxygen of the bovine serum albumin aminoacid residue amounts 580 kJ/mol. The 3-D bioconstructions based on MWCNTs and SWCNTs becomes overgrown with the cells (fibroblasts) over the course of 72 hours. The samples based on the both types of CNTs are not toxic for the cells and don't change its normal composition and structure. Thus the 3-D bioconstructions that are nanostructured by the pulsed and continuous laser radiation can be applied as implant materials for the recovery of the connecting tissues of the living body.

In this paper, we present an original concept of plasmonic-related instrumentation platform dedicated to diagnostic biosensing tests out of the laboratory. The developed instrumental platform includes both disposable one-use microfluidic affinity biochip and compact optical readout device for biochip monitoring involving mobile Internet devices for data processing and communication. The biochip includes both microfluidic and optical coupling structures formed into a single plastic slab. The microfluidic path of the biochip operates in passive capillary pumping mode. In the proof-of-concept prototype, we address specifically the sensing format involving Surface Plasmon Resonance phenomenon. The biochip is plugged in the readout device without the use of an index matching fluid. An essential advantage of the developed biochip is that its implementation involves conventional hot embossing and thin film deposition process, perfectly suited for mass production of low-cost microfluidic biochip for biochemical applications.

In this work, two plasmonic optical fiber sensor configurations are used for protein sensing and their relative performances in terms of limit of detection and sensitivity are compared. The first configuration consists in unclad 200 μm optical fibers that produce a broadband resonance in the visible wavelength range around 650 nm while the second configuration makes use of multiple narrowband resonances produced in the C+L bands with weakly tilted fiber Bragg gratings photo-inscribed in telecommunication-grade single-mode optical fibers. In both cases, the sensitive regions are surrounded by a ~50 nm gold layer so that the evanescent wave can excite a surface plasmon polariton at the metalsurrounding medium interface. Both configurations are used to sense green fluorescent proteins. Our experimental results demonstrate that the two sensor configurations present a complementary measurement dynamics as a function of the investigated concentration in the range 10^-12 – 10^-7 g/ml. We attribute this difference of sensitivity to the difference of penetration depth of the evanescent wave in the surrounding medium, which is proportional to the light wavelength.

The ageing population faces today an increase of chronic diseases such as rheumatism/arthritis, cancer and cardio vascular diseases for which appropriate treatments based on a diagnosis at an early-stage of the disease are required. Some imaging techniques are already available in order to get structural information. Within the non-invasive group, ultrasound images are common in these fields of medicine. However, there is a need for a point-of-care device for imaging smaller structures such as blood vessels that cannot be observed with purely ultrasound based devices. Photoacoustics proved to be an attractive candidate. This novel imaging technique combines pulsed laser light for excitation of tissues and an ultrasound transducer as a receptor. Introduction of this technique into the clinic requires to drastically shrink the size and cost of the expensive and bulky nanosecond lasers generally used for light emission. In that context, demonstration of ultra-short pulse emission with highly efficient laser diodes in the near-infrared range has been performed by Quantel, France. A multi-wavelength laser source as small as a hand emitted more than 1 mJ per wavelength with four different wavelengths available in pulses of about 90 ns. Such a laser source can be integrated into high sensitivity photoacoustic handheld systems due to their outstanding electrical-to-optical efficiency of about 25 %. Further work continues to decrease the pulse length as low as 40 ns while increasing the pulse energy to 2 mJ.

The research and development of personalized medical treatments is increasing steadily fostered by its large societal impact. The ability of non-spherical gold nanoparticles to locally and efficiently release heat when irradiated in Near Infrared (NIR) wavelength region is a promising tool for photothermal medical therapies. In the present work, stable inks containing PEGylated gold nanostars (GNS) were obtained and inkjet-printed on a pigment coated paper substrate. Significant photothermal effect of the printed patterns was observed under Near Infrared (NIR) excitation of the Localized Surface Plasmon Resonance (LSPR) of the GNS. These preliminary results support, in perspective, the application of printed GNS patterns for thermal medical treatments either by direct localized heating, or by temperature triggered drug release.

Atherosclerosis is a narrowing of the arteries caused by an increase of atheromatous plaque: material formed by macrophage cells containing cholesterol and fatty acids, calcium and a variable amount of fibrous connective tissue. The elation between vulnerable plaques and cardiovascular events can be determined using plaque biomarkers. In this work, atherosclerotic plaques stained with different molar ratios of europium, in a potential plaque biomarker, europium tetracycline complex, were studied by fluorescence microscopy. The tetracycline antibiotic used was chlortetracycline. The growth of atherosclerotic plaque was followed during 60 days in New Zealand rabbits divided in two groups: an experimental group (EG), with nine animals and a control group (CG) with three animals. The animals in the EG received a diet with 1% of cholesterol and the animals of GC received a normal diet. The aortic arch of the animals with 60 days were cut in the vertical plane in 6 μm thick slices, which were mounted on glass slides and stained with hematoxylin an eosin and europium chlortetracycline complex (EuCTc). The fluorescence images were obtained exciting the EuCTc absorption band with a filter cube D (BP 355 – 425) and the emission was collected with a LP 470 suppression filter. Light intensity, detector gain and acquisition time were fixed for comparisons. The 20× magnified images were collected with 12 bit (or 4096 gray tones) resolution. The mean value of gray scale for each molar ratio of EuCTc was different, indicating that the complex interacts with the components of atherosclerotic plaque and the best molar ratio was 1.5 EuCTc. These results indicate the potential use of the EuCTc biomarker for atherosclerotic plaque characterization.

Uniform illumination of a working field is very important in optical systems of confocal microscopy and various implementations of fluorescence microscopy like TIR, SSIM, STORM, PALM to enhance performance of these laser-based research techniques. Widely used TEM₀₀ laser sources are characterized by essentially non-uniform Gaussian intensity profile which leads usually to non-uniform intensity distribution in a microscope working field or in a field of microlenses array of a confocal microscope optical system, this non-uniform illumination results in instability of measuring procedure and reducing precision of quantitative measurements. Therefore transformation of typical Gaussian distribution of a TEM₀₀ laser to flat-top (top hat) profile is an actual technical task, it is solved by applying beam shaping optics. Due to high demands to optical image quality the mentioned techniques have specific requirements to a uniform laser beam: flatness of phase front and extended depth of field, - from this point of view the microscopy techniques are similar to holography and interferometry. There are different refractive and diffractive beam shaping approaches used in laser industrial and scientific applications, but only few of them are capable to fulfil the optimum conditions for beam quality required in discussed microscopy techniques. We suggest applying refractive field mapping beam shapers πShaper, which operational principle presumes almost lossless transformation of Gaussian to flat-top beam with flatness of output wavefront, conserving of beam consistency, providing collimated low divergent output beam, high transmittance, extended depth of field, negligible wave aberration, and achromatic design provides capability to work with several lasers with different wavelengths simultaneously. The main function of a beam shaper is transformation of laser intensity profile, further beam transformation to provide optimum for a particular technique spot size and shape has to be realized by an imaging optical system which can include microscope objectives and tube lenses. This paper will describe design basics of refractive beam shapers and optical layouts of their applying in microscopy systems. Examples of real implementations and experimental results will be presented as well.

Skin health characterized by a system of water and lipids in Stratum Corneum provide protection from harmful external elements and prevent trans-epidermal water loss. Skin hydration (moisture) and sebum (skin surface lipids) are considered to be important factors in skin health; a right balance between these components is an indication of skin health and plays a central role in protecting and preserving skin integrity. In this manuscript we present an infrared spectroscopic method for simultaneous and quantitative measurement of skin hydration and sebum levels utilizing differential detection with three wavelengths 1720, 1750, and 1770 nm, corresponding to the lipid vibrational bands that lie “in between” the prominent water absorption bands. The skin sebum and hydration values on the forehead under natural conditions and its variations to external stimuli were measured using our experimental set-up. The experimental results obtained with the optical set-up show good correlation with the results obtained with the commercially available instruments Corneometer and Sebumeter.

Neonatal hemorrhagic stroke (NHS) is a major problem of future generation’s health due to the high rate of death and cognitive disability of newborns after NHS. The incidence of NHS in neonates cannot be predicted by standard diagnostic methods. Therefore, the identification of prognostic markers of NHS is crucial. There is evidence that stress-related alterations of cerebral blood flow (CBF) may contribute to NHS. Here, we assessed the stroke-associated CBF abnormalities for high prognosis of NHS using a new model of NHS induced by sound stress in the pre- and post-stroke state. With this aim, we used interdisciplinary methods such as a histological assay of brain tissues, laser speckle contrast imaging and Doppler coherent tomography to monitor cerebral circulation. Our results suggest that the venous stasis with such symptoms as progressive relaxation of cerebral veins, decrease the velocity of blood flow in them are prognostic markers for a risk of NHS and are an informative platform for a future study of corrections of cerebral venous circulatory disturbance related to NHS.

Quantitative Mueller polarimetry optically characterizes a medium and is reflected upon by the ultrastructural changes in it. Tissue morphology changes occur during advent of diseases like cancer neoplasia. This alters the Mueller matrix characterizing the tissue as an optical element. The nucleus size undergoes an approximate doubling during the development of cancer. Cell crowding during cancer increases the number density of the nuclei per unit volume. Modeling the cell nuclei as main scattering centers, a systematic computational study on how Mueller matrix elements vary for an increase in scatterer size and number density is performed. Simulation on polarized light transport of wavelength 633nm through a slab of size 3 mm comprising of spherical scatterers in a medium of refractive index 1.33 is carried out. Light propagation is modeled using Monte Carlo method and meridian plane method is adopted for tracking the polarization state change. The stokes vector of the outgoing light is tracked to calculate the Mueller matrix images of the light backscattered from the slab. The Mueller matrix elements as well as depolarization factors are derived. The depolarization index increases with scatterer size. Along with nucleus size, change in the cell number density is also expected in the different stages of the cancer growth. Volume fraction of the scatterers in medium is varied as an indicator of this number density change. Behavior of Mueller matrix with respect to change in scattering coefficient due to variation in scatterer size and volume fraction is studied. It is observed that the depolarization index derived from Mueller matrix has selective discrimination towards the change in scattering coefficient caused due to size change and volume fraction change respectively.

The aim of the present work was to study energy metabolism in human cervical carcinoma (HeLa) cells in vitro and in vivo using two-photon FLIM. Cellular metabolism was examined by monitoring of the fluorescence lifetimes of free and protein-bound forms of NAD(P)H and FAD and their relative contributions. Two-photon fluorescence and second harmonic generation microscopy as well as standard histopathology with hematoxylin and eosin were used to characterize tissue structure. Cellular metabolism was analyzed in cancer cells co-cultured with human fibroblasts and in tumor xenografts transplanted to nude mice. In the HeLa-huFB co-culture we observed a metabolic shift from OXPHOS toward glycolysis in cancer cells, and from glycolysis to OXPHOS in fibroblasts, starting from Day 2 of co-culturing. In the tumor tissue we detected metabolic heterogeneity with more glycolytic metabolism of cancer cells in the stroma-rich zones. The results of the study are of a great importance for understanding metabolic behavior of tumors and for development of anticancer drugs targeted to metabolic pathways.

A novel direct spectrophotometric method for quantitative determination of Oxiphore® drug substance (synthetic polyhydroquinone complex) in food supplements is developed. Absorption spectra of Oxiphore® water solutions in the ultraviolet region are presented. Samples preparation procedures and mathematical methods of spectra post-analytical procession are discussed. Basic characteristics of the automatic CCD-based UV spectrophotometer and special software implementing the developed method are described. The results of the trials of the developed method and software are analyzed: the error of determination for Oxiphore® concentration in water solutions of the isolated substance and singlecomponent food supplements did not exceed 15% (average error was 7…10%).

5-aminolevulinic-acid-(5-ALA)-induced protoporphyrin IX (PpIX) fluorescence may be used to improve stereotactic brain tumor biopsies. In this study, the sensitivity of PpIX-based tumor detection has been investigated for two potential excitation wavelengths (405 nm, 633 nm). Using a 200 μm fiber in contact with semi-infinite optical phantoms containing ink and Lipovenös, PpIX detection limits of 4.0 nM and 200 nM (relating to 1 mW excitation power) were determined for 405 nm and 633 nm excitation, respectively. Hence, typical PpIX concentrations in glioblastomas of a few μM should be well detectable with both wavelengths. Additionally, blood layers of selected thicknesses were placed between fiber and phantom. Red excitation was shown to be considerably less affected by blood interference: A 50 μm blood layer, for instance, blocked the 405- nm-excited fluorescence completely, but reduced the 633-nm-excited signal by less than 50%. Ray tracing simulations demonstrated that – without blood layer – the sensitivity advantage of 405 nm rises for decreasing fluorescent volume from 50-fold to a maximum of 100-fold. However, at a tumor volume of 1 mm3, which is a typical biopsy sample size, the 633-nm-excited fluorescence signal is only reduced by about 10%. Further simulations revealed that with increasing fiber-tumor distance, the signal drops faster for 405 nm. This reduces the risk of detecting tumor tissue outside the needle’s coverage, but diminishes the overlap between optically and mechanically sampled volumes. While 405 nm generally offers a higher sensitivity, 633 nm is more sensitive to distant tumors and considerably superior in case of blood-covered tumor tissue.

Disinfection routines are important in all clinical applications. The uprising problem of antibiotic resistance has driven major research efforts towards alternative disinfection approaches, involving light-based solutions. Pseudomonas aeruginosa (P. aeruginosa) is a common bacterium that can cause skin, soft tissue, lungs, kidney and urinary tract infections. Moreover, it can be found on and in medical equipment causing often cross infections in hospitals. The objective of this study was to test the efficiency, of two different light-based disinfection treatments, namely UVB and UVC irradiation, on P. aeruginosa biofilms at different growth stages. In our experiments a new type of UV light emitting diodes (LEDs) were used to deliver UV irradiation on the biofilms, in the UVB (296nm) and UVC (266nm) region. The killing rate was studied as a function of dose for 24h grown biofilms. The dose was ramped from 72J/m2 to 10000J/m2. It was shown that UVB irradiation was more effective than UVC irradiation in inactivating P. aeruginosa biofilms. No colony forming units (CFU) were observed for the UVB treated biofilms when the dose was 10000 J/m2 (CFU in control sample: 7.5 x 10⁴). UVB irradiation at a dose of 20000J/m2 on mature biofilms (72h grown) resulted in a 3.9 log killing efficacy. The fact that the wavelength of 296nm exists in daylight and has such disinfection ability on biofilms gives new perspectives for applications within disinfection at hospitals.

We have analyzed here low coherence spectroscopic data by multifractal analysis for obtaining tissue multifratality in depth wise index distribution. Essentially, in this method, a spectral domain interference pattern is recorded in a common path interferometer with a broad band source operated in back scattering mode. The recorded interference spectrum is subjected to Fourier domain processing to compute depth wise index distribution with a resolution of the order of one micron. The experimental set-up was validated, initially, by verifying depths of mica sheet layers and diameter of polystyrene microspheres and later, it was used for assessment of depth wise index (RI) distribution of cervical tissue slices. The structures of cervical tissues at different stages of cancer change rapidly which are manifested in the RI distribution and in turn, are encoded as multi-resolution information. This information can, effectively, be extracted by using multifractal detrended fluctuation analysis (MFDFA), where, multifractal parameters such as Hurst exponent h(q = 2) and width of singularity spectrum (Δα) show definitive change as cancer progresses from grade I to grade II. Moreover, the depth distribution of RI exhibited stronger multifractality (increased Δα) and considerably weakened correlations (decreased h(q = 2)) for tissues with higher pathological grades. Therefore, the technique of low coherence back-scattered (LCBS) interferometry bears a promise of using depth distribution of tissue refractive index and MFDFA analysis appears as a label free biomarker to detect cancer at early.

Antioxidants, reactive oxygen species and other radicals are involved in oxidative stress phenomena. There are many available methods to quantify the progress of the process and evidence its existence for each stage of oxidative damage process which can be classified into direct and indirect approaches. The technic of experimental determination of optical characteristics stability of antioxidant solutions with spectroscopic method was offered in this work as a direct method for investigation of color stability in time and fluorescence behavior of antioxidant solutions. Coenzyme Q10, or ubiquinone, in the form of oil and alcohol solutions was chosen as an antioxidant of research. The experimental results connected with the optical density and color characteristics confirm the possibility of applying the clinical diagnosis method of estimation redox potential. Also, future prospects, including application of this technic in pharmaceutical analysis for quality control, were discussed.

Fluorescent organic dyes solutions are used for non-contact measurement of the millimeter wave absorption in liquids simulating biological tissue. There is still not any certain idea of the physical mechanism describing this process despite the widespread technology of microwave radiation in the food industry, biotechnology and medicine. For creating adequate physical model one requires an accurate command of knowledge concerning to the relation between millimeter waves and irradiated object. There were three H-bonded liquids selected as the samples with different coefficients of absorption in the millimeter range like water (strong absorption), glycerol (medium absorption) and ethylene glycol (light absorption). The measurements showed that the greatest response to the action of microwaves occurs for glycerol solutions: R6G (building-up luminescence) and RC (fading luminescence). For aqueous solutions the signal is lower due to lower quantum efficiency of luminescence, and for ethylene glycol — due to the low absorption of microwaves. In the area of exposure a local increase of temperature was estimated. For aqueous solutions of both dyes the maximum temperature increase is about 7° C caused with millimeter waves absorption, which coincides with the direct radio physical measurements and confirmed by theoretical calculations. However, for glycerol solution R6G temperature equivalent for building-up luminescence is around 9° C, and for the solution of ethylene glycol it’s about 15°. It is assumed the possibility of non-thermal effect of microwaves on the different processes and substances. The application of this non-contact temperature sensing is a simple and novel method to detect temperature change in small biological objects.

In this study we demonstrate the capability of cross-polarization optical coherence tomography (CP OCT) to assess collagen and elastin fibers condition in atherosclerotic plaques basing on ratio of the OCT signal levels in cross- and со- polarizations. We consider the depolarization factor (DF) and the effective birefringence (Δn) as quantitative characteristics of CP OCT images. We revealed that calculation of both DF and Δn in the region of interest (fibrous cap) yields a statistically significant difference between stable and unstable plaques (0.46±0.21 vs 0.09±0.04 for IDF; (4.7±1.0)•10^–4 vs (2.5±0.7)•10^–4 for Δn; p<0.05). In parallel with CP OCT we used the nonlinear microscopy for analysis of thin cross-section of atherosclerotic plaque, revealing the different average isotropy index of collagen and elastin fibers for stable and unstable plaques (0.30 ± 0.10 vs 0.70 ± 0.08; р<0.001). The proposed approach for quantitative assessment of CP OCT images allows cross-scattering and birefringence characterization of stable and unstable atherosclerotic plaques.

Optical properties of mesoporous TiO₂ scatters with average diameters of 20 nm, 150 nm, 300 nm, and 500 nm are measured by optical coherence tomography system at the wavelength of 853 nm. The scattering coefficient can be estimated by utilizing the extend Huygens Fresnel theory with consideration of single and multiple scattering effects, and the g factors are calculated based on the Mie scattering model. After fitting algorithm, by increasing the sizes of mesoporous TiO₂ scatters from 20, 150, 300, to 500 nm, the scattering coefficient could also be extracted as from 13.5±0.6, 16.5±0.6, to 20.2±0.6 mm-1, and then decreased to 18.5±0.6 mm^-1, which are accompanied with the enhancement of multiple-scattering contribution from 5%, 10%, to 40%, and then decreased to 20%, respectively.

Here we report our investigations of the formation conditions and photophysical properties of complexes between luminescent semiconducting nanoparticles (quantum dots, QDs) and the photosensitizer chlorin e6, which is widely used for the photodynamic therapy. In our complexes, bovine serum albumin (BSA), the most abundant protein in blood serum, was used as a linker between QDs and chlorin e6 molecules. The influence of BSA on the optical properties of Ce6 and QDs in complexes was properly examined using spectral-luminescent methods. It was found that BSA passivated QD surface and substantially QD quantum yield of luminescence was increased. In addition, BSA prevented the aggregation of chlorin e6 molecules in complexes with QDs. We demonstrated that the use of BSA as a linker allows to create functional QD-chlorin e6 complexes with effective photoexcitation energy transfer from QDs to the molecules.

The pinhole plays an important role in confocal laser scanning microscopy (CLSM) for adaptive optics (AO) as well as in imaging, where the size of the pinhole denotes a trade-off between out-of-focus rejection and wavefront distortion. This contribution proposes an AO system for a commercial CLSM with an adjustable square pinhole to cope with such a trade-off. The proposed adjustable pinhole enables to calibrate the AO system and to evaluate the imaging performance. Experimental results with fluorescence beads on the coverslip and at a depth of 40 μm in the human hepatocellular carcinoma cell spheroid demonstrate that the proposed AO system can improve the image quality by the proposed calibration method. The proposed pinhole intensity ratio also indicates the image improvement by the AO correction in intensity as well as resolution.

Raman micro-spectroscopy is an optoelectronic technique that can be used to evaluate the chemical composition of biological samples and has been shown to be a powerful diagnostic tool for the investigation of various cancer related diseases including bladder, breast, and cervical cancer. Raman scattering is an inherently weak process with approximately 1 in 107 photons undergoing scattering and for this reason, noise from the recording system can have a significant impact on the quality of the signal, and its suitability for diagnostic classification. The main sources of noise in the recorded signal are shot noise, CCD dark current, and CCD readout noise. Shot noise results from the low signal photon count while dark current results from thermally generated electrons in the semiconductor pixels. Both of these noise sources are time dependent; readout noise is time independent but is inherent in each individual recording and results in the fundamental limit of measurement, arising from the internal electronics of the camera. In this paper, each of the aforementioned noise sources are analysed in isolation, and used to experimentally validate a mathematical model. This model is then used to simulate spectra that might be acquired under various experimental conditions including the use of different cameras, different source wavelength, and power etc. Simulated noisy datasets of T24 and RT112 cell line spectra are generated based on true cell Raman spectrum irradiance values (recorded using very long exposure times) and the addition of simulated noise. These datasets are then input to multivariate classification using Principal Components Analysis and Linear Discriminant Analysis. This method enables an investigation into the effect of noise on the sensitivity and specificity of Raman based classification under various experimental conditions and using different equipment.

This paper presents the results of thermal and optical modeling of "blackened" tips (fiber-optic thermal converter) with different structures: film and volumetric. Film converter is created by laser radiation action on a cork or paper and it is a one-step process. As a result, a carbonized cork or paper adhered to the distal end of the optical fiber absorbs light that leads to heating of the distal end of the optical fiber. We considered the peculiarities of volumetric converters formed by sintering (second step) of the target material transferred to the tip, at irradiating the target with laser radiation (first step). We investigated the interaction between 980 nm laser radiation and converters in the air and water. As a result of experiments and modeling, it was obtain, that converter temperature and power of converter destruction depend on the environment in which it is placed. We found that film converter in the air at average power of laser radiation of 0.30±0.05 W is heated to 900±50°С and destructed, and volumetric converter in the air at average power of laser radiation of 1.0±0.1 W is heated to 1000±50°С and destructed at reaching of 4.0±0.1 W only. We found that film converter in the water at average power of laser radiation of 1.0±0.1 W is heated to 550±50°С and destructed at reaching of 4.0±0.1 W only. Volumetric converter at average power of laser radiation of4.0±0.1 W is heated to 450±50°С and is not destructed up to 7.5±0.1 W, it is heated to 500±50°С in this case. Thus, volumetric converter is more resistant to action of laser heating.

In this study we demonstrate a comparative analysis of blood serum and normal human skin by Raman spectroscopy with application of different spectroscopic equipment. For serum analysis we measure a total concentration of proteins and compared it with intensity of 1002 cm^-1 Raman peak. Standard deviation for protein control in blood serum differed from 7.4% to 19% for different spectroscopic setups. For human skin control we used three Raman peaks near 1340, 1450 and 1650 cm^-1. Measurements of different skin samples were analyzed on the phase plane to find areas corresponding to the normal skin. Taking into account the different sensitivities of the detected signal with different detectors in the spectral range 810-950 nm we calculated correction coefficients allowed for making comparison of spectral measurements made on different spectrometers with ranging not exceeding 21%.

Optical coherence tomography (OCT) is usually employed for the measurement of tumor topology, which reflects structural changes of a tissue. We investigated the possibility of OCT in detecting changes using a computer texture analysis method based on Haralick texture features, fractal dimension and the complex directional field method from different tissues. These features were used to identify special spatial characteristics, which differ healthy tissue from various skin cancers in cross-section OCT images (B-scans). Speckle reduction is an important pre-processing stage for OCT image processing. In this paper, an interval type-II fuzzy anisotropic diffusion algorithm for speckle noise reduction in OCT images was used. The Haralick texture feature set includes contrast, correlation, energy, and homogeneity evaluated in different directions. A box-counting method is applied to compute fractal dimension of investigated tissues. Additionally, we used the complex directional field calculated by the local gradient methodology to increase of the assessment quality of the diagnosis method. The complex directional field (as well as the “classical” directional field) can help describe an image as set of directions. Considering to a fact that malignant tissue grows anisotropically, some principal grooves may be observed on dermoscopic images, which mean possible existence of principal directions on OCT images. Our results suggest that described texture features may provide useful information to differentiate pathological from healthy patients. The problem of recognition melanoma from nevi is decided in this work due to the big quantity of experimental data (143 OCT-images include tumors as Basal Cell Carcinoma (BCC), Malignant Melanoma (MM) and Nevi). We have sensitivity about 90% and specificity about 85%. Further research is warranted to determine how this approach may be used to select the regions of interest automatically.

Photoplethysmography (PPG) technology is widely used in wearable heart pulse rate monitoring. It might reveal the potential risks of heart condition and cardiopulmonary function by detecting the cardiac rhythms in physical exercise. However the quality of wrist photoelectric signal is very sensitive to motion artifact since the thicker tissues and the fewer amount of capillaries. Therefore, motion artifact is the major factor that impede the heart rate measurement in the high intensity exercising. One accelerometer and three channels of light with different wavelengths are used in this research to analyze the coupled form of motion artifact. A novel approach is proposed to separate the pulse signal from motion artifact by exploiting their mixing ratio in different optical paths. There are four major steps of our method: preprocessing, motion artifact estimation, adaptive filtering and heart rate calculation. Five healthy young men are participated in the experiment. The speeder in the treadmill is configured as 12km/h, and all subjects would run for 3-10 minutes by swinging the arms naturally. The final result is compared with chest strap. The average of mean square error (MSE) is less than 3 beats per minute (BPM/min). Proposed method performed well in intense physical exercise and shows the great robustness to individuals with different running style and posture.

The experimental results presented in this study focused on the study of biological processes caused by exposure to the coating layers of the laser green light seed (λ = 532 nm) range for the larch, violet (λ = 405 nm) and red (λ = 640 nm) for spruce. Spend a series of experiments to study the dependence of crop seed quality (spruce and larch from the pine family) from exposure to laser radiation under different conditions. In all the analyzed groups studied seed germination and growth of seedlings exposed to laser exposure, compared with the control group. The results showed that the higher percentage of germination than seeds of the control group.

Experimental results presented in this study tends to explore viruses in the water for their further decontamination under the influence of laser radiation (λ=220-390 nm). Conducted a series of experiments to study the dependence of water quality from the effects of laser radiation. Correlation between degree of survival of viruses and power density. The results showed that all the analyzed samples of water is clearing from bacteria to 98%. Preliminary tests of the prototype laboratory system UFOVI has opened up new opportunities for water sterilizing.

BioMérieux is automating the microbiology laboratory in order to reduce cost (less manpower and consumables), to improve performance (increased sensitivity, machine algorithms) and to gain traceability through optimization of the clinical laboratory workflow. In this study, we evaluate the potential of Hyperspectral imaging (HSI) as a substitute to human visual observation when performing the task of microbiological culture interpretation. Microbial colonies from 19 strains subcategorized in 6 chromogenic classes were analyzed after a 24h-growth on a chromogenic culture medium (chromID® CPS Elite, bioMérieux, France). The HSI analysis was performed in the VNIR region (400-900 nm) using a linescan configuration. Using algorithms relying on Linear Spectral Unmixing, and using exclusively Diffuse Reflectance Spectra (DRS) as input data, we report interclass classification accuracies of 100% using a fully automatable approach and no use of morphological information. In order to eventually simplify the instrument, the performance of degraded DRS was also evaluated using only the most discriminant 14 spectral channels (a model for a multispectral approach) or 3 channels (model of a RGB image). The overall classification performance remains unchanged for our multispectral model but is degraded for the predicted RGB model, hints that a multispectral solution might bring the answer for an improved colony recognition.