МЕТОД БЕСКОНТАКТНОЙ ФОТОЛЮМИНЕСЦЕНТНОЙ ДИАГНОСТИКИ СОСТОЯНИЯ ФИБРОЗНОЙ ОБОЛОЧКИ ГЛАЗА

Non-contact optical diagnostics of structural disorders of the eye has a number of advantages: high speed, accuracy and a large range of parameters available for analysis. The paper presents the results of studies of the photoluminescence of the fibrous tunic of the eye, excited by polarized light, depending on the intraocular pressure. In the experiments, isolated de-epithelized eyes of the rabbit were used, inside of which pressure up to 50 mm Hg was artificially created. Under these conditions, the cornea and sclera were illuminated with linearly polarized light at wavelengths of 250, 350 and 450 nm, exciting photoluminescence in the wavelength range up to 700 nm. Cross and co-polarized photoluminescence spectra excited by linearly polarized light were obtained. It has been established that, when excited by polarized light, the photoluminescence of the cornea is partially polarized. Depending on the wavelength of the photoluminescence, the degree of polarization varies from 0.2 to 0.35. It is shown that the degree of polarization of the photoluminescence of the cornea of the eye upon excitation by linearly polarized light can be used as a measurable parameter for assessing the physiological state of the eye. It is shown that the photoluminescence spectrum consists of two bands with maxima near 460-470 and 430-440 nm. These bands are assigned, respectively, to pyridinnucleotides and glycosylated collagen. A significant contribution can be made by the epithelium of the eye, which contains riboflavin with characteristic absorption bands near 450 and 365 nm. When excited at 450 nm, the photoluminescence maximum is located near 540 nm, which corresponds to the spectrum of fluorophores in the endothelium and epithelium. The spectrum of photoluminescence upon excitation at a wavelength of 250 nm can be attributed to tryptophan located in the intraocular lens.Бесконтактная оптическая диагностика структурных нарушений глаза обладает рядом преимуществ: высокая скорость, точность и большой спектр параметров, доступных для анализа. В работе представлены результаты исследований фотолюминесценции фиброзной оболочки глаза, возбуждаемой поляризованным светом, в зависимости от внутриглазного давления. В эксперименте применяли деэпителизированные глаза кролика с искусственно повышенным офтальмотонусом до 50 мм рт.ст. При этом склеру и роговицу освещали линейно поляризованным светом на длинах волн 250, 350 и 450 нм, возбуждая фотолюминесценцию в диапазоне длин волн до 700 нм. Были получены кои кросс-поляризованные спектры фотолюминесценции, возбуждаемые линейно поляризованным светом. При возбуждении поляризованным светом фотолюминесценция роговицы оказалась частично поляризованной. В зависимости от длины волны фотолюминесценции степень поляризации изменяется от 0,2 до 0,35. Показано, что степень поляризации фотолюминесценции роговицы глазапривозбуждении линейно поляризованным светом можно рассматривать в качестве измеряемого параметра для оценки состояния внутриглазного давления. Показано, что спектр фотолюминесценции состоит из двух полос с максимумами вблизи 460-470 и 430-440 нм. Эти полосы отнесены, соответственно, к пиридиннуклеотидамигликозилированному коллагену. Существенный вклад оказывает эпителий глаза, в котором содержится рибофлавин с полосами поглощения вблизи длин волн 450 и 365 нм. При возбуждении на длине волны 450 нм максимум фотолюминесценции расположен вблизи 540 нм, что соответствует спектру флуорофоров в эндотелии и эпителии. Спектр фотолюминесценции при возбуждении на длине волны 250 нм можно приписать триптофану, находящемуся в хрусталике глаза

Incoherent wavefront reconstruction by a retroemission device

This Letter addresses wavefront reconstruction by a retroemission device (REM). REM represents a lenslet array mounted on a substrate made of photoluminescent optical material, such as a polymer film impregnated with upconversion nanoparticles. An excitation light wavefront incident on the REM was sampled by the lenslet array piece-wise. Each wavelet at the lenslet aperture was converged into a voxel in the substrate, with its coordinates encoding the angle of incidence and curvature of the wavelet. Photoluminescence excited in the voxel was radiated isotropically, its back-propagating fraction was captured by the lenslet and transformed into a back-propagating wavelet, which contributed to reproduction of the entire incident wavefront with some fidelity. We experimentally proved the wavefront reconstruction based on REM, and present its theoretical model based on a Fresnel-Kirchhoff approximation.4 page(s

<i>In vitro и in vivo</i> photodynamic therapy of solid tumors with a combination of riboflavin and upconversion nanoparticles

Rationale: Riboflavin (vitamin B2) is one of the most promising agents for photodynamic therapy (PDT). However, its use is limited by the excitation in the ultraviolet (UV) and visible spectral ranges and, as a result, by a small penetration into biological tissue not exceeding a few millimeters. This problem could be solved by approaches ensuring excitation of riboflavin molecules within tumor tissues by infrared (IR) light. Upconversion nanoparticles (UCNPs) can be potentially considered as mediators able to effectively convert the exciting radiation of the near IR range, penetrating into biological tissue to a 3 cm depth, into the photoluminescence in the UV and visible spectral ranges.Aim: To evaluate the efficacy of UCNPs for IR-mediated riboflavin activation in the depth of tumor tissue during PDT. Materials and methods: The water-soluble riboflavin flavin mononucleotide (FMN, Pharmstandard-UfaVITA, Russia) was used as a photosensitizer in in vitro and in vivo experiments. The in vitro experiments were performed on human breast adenocarcinoma SK-BR-3, human glioblastoma U-87 MG, and rat glioma C6 cell lines. Lewis lung carcinoma (LLC) inoculated to hybrid BDF1 mice was used as a model to demonstrate the delivery of FMN to the tumor. UCNPs with a core/shell structure [NaYF4:Yb3+, Tm3+/NaYF4] were used for photoactivation of FMN in vivo. PDT based on FMN, UCNPs and laser radiation 975 nm (IR) was performed on mouse xenografts of human breast adenocarcinoma SKBR-3.Results: We were able to show that FMN could act as an effective in vitro photosensitizer for SK-BR-3, U-87 MG, and C6 cell lines. FMN IC50 values for glioma cells were ~30 μM, and for SK-BR-3 cell line ~50 μM (24 h incubation, irradiation 4.2 J/cm2). In the LLC model, the appropriate concentration of FMN (30 μM and above) can be achieved in the tumor as a result of systemic administration of FMN (at 2 and 24 hours after injection). The effect of PDT using near IR light for UCNP-mediated excitation of FMN was demonstrated in mouse xenografts SKBR-3, with the tumor growth inhibition of 90±5%.Conclusion: The study has demonstrated the possibility to use riboflavin (vitamin B2) as a photosensitizer for PDT. The photoexcitation of FMN via the anti-Stokes photoluminescence of UCNPs allows for implementation of the PDT technique with the near IR spectral range

METHOD OF NON-CONTACT PHOTOLUMINESCENT DIAGNOSTICS OF THE EYE FIBROUS TUNIC CONDITION

Non-contact optical diagnostics of structural disorders of the eye has a number of advantages: high speed, accuracy and a large range of parameters available for analysis. The paper presents the results of studies of the photoluminescence of the fibrous tunic of the eye, excited by polarized light, depending on the intraocular pressure. In the experiments, isolated de-epithelized eyes of the rabbit were used, inside of which pressure up to 50 mm Hg was artificially created. Under these conditions, the cornea and sclera were illuminated with linearly polarized light at wavelengths of 250, 350 and 450 nm, exciting photoluminescence in the wavelength range up to 700 nm. Cross and co-polarized photoluminescence spectra excited by linearly polarized light were obtained. It has been established that, when excited by polarized light, the photoluminescence of the cornea is partially polarized. Depending on the wavelength of the photoluminescence, the degree of polarization varies from 0.2 to 0.35. It is shown that the degree of polarization of the photoluminescence of the cornea of the eye upon excitation by linearly polarized light can be used as a measurable parameter for assessing the physiological state of the eye. It is shown that the photoluminescence spectrum consists of two bands with maxima near 460-470 and 430-440 nm. These bands are assigned, respectively, to pyridinnucleotides and glycosylated collagen. A significant contribution can be made by the epithelium of the eye, which contains riboflavin with characteristic absorption bands near 450 and 365 nm. When excited at 450 nm, the photoluminescence maximum is located near 540 nm, which corresponds to the spectrum of fluorophores in the endothelium and epithelium. The spectrum of photoluminescence upon excitation at a wavelength of 250 nm can be attributed to tryptophan located in the intraocular lens

Near-infrared photopolymerization assisted by upconversion nanophosphors for biomedical applications

We present the concept and the experimental demonstration of near-infrared photopolymerization assisted by specially designed upconversion nanophosphors. The principle of this technique is based on conversion of 980 nm laser irradiation to ultraviolet photons subsequently absorbed by photoinitiator. The nonlinearity of upconversion allows for activation of the process locally in the laser beam waist. This approach enables precise fabrication of 3D constructs directly in the volume of photocurable composition. Furthermore, the presented technique is suitable for polymerization of a wide range of photocurable resins as well as gelation of hydrogels for biomedical applications

Near-infrared photopolymerization assisted by upconversion nanophosphors for biomedical applications

We present the concept and the experimental demonstration of near-infrared photopolymerization assisted by specially designed upconversion nanophosphors. The principle of this technique is based on conversion of 980 nm laser irradiation to ultraviolet photons subsequently absorbed by photoinitiator. The nonlinearity of upconversion allows for activation of the process locally in the laser beam waist. This approach enables precise fabrication of 3D constructs directly in the volume of photocurable composition. Furthermore, the presented technique is suitable for polymerization of a wide range of photocurable resins as well as gelation of hydrogels for biomedical applications

LUMINESCENCE DIAGNOSTICS OF TUMORS WITH UPCONVERSION NANOPARTICLES

Background: To improve quality of surgery in oncology, it is necessary to completely remove the tumor, including its metastases, to minimize injury to normal tissues and to reduce duration of an intervention. Modern methods of detection based on radiological computerized tomography and magnetic resonance imaging can identify a tumor after its volume has become big enough, i.e. it contains more than 10 billion cells. Therefore, an improvement of sensitivity and resolution ability of diagnostic tools to identify early stages of malignant neoplasms seems of utmost importance. Aim: To demonstrate the potential of a new class of anti-Stokes luminescence nanoparticles for deep optical imaging with high contrast of malignant tumors. Materials and methods: Upconversion nanoparticles with narrow dispersion and a  size of 70 to 80  nm, with a  core/shell structure of NaYF4:Yb3+:Tm3+/NaYF4 were used in the study. The nanoparticles have an intensive band of anti-Stokes photoluminescence at a wavelength of 800  nm under irradiation with a  wavelength of 975  nm (both wavelengths are within the transparency window for biological tissues). The conversion coefficient of the excitation radiation into the anti-Stokes luminescence was 9%. To increase the time during which nanoparticles can circulate in blood flow of small animals, the nanoparticles were covered by a  biocompatible amphiphilic polymer shell. As a  tumor model we used Lewis epidermoid carcinoma transfected to mice. Results: We were able to obtain stable water colloids of nanoparticles covered with amphiphilic polymer that could preserve their initial size at least for one month. The use of upconversion nanoparticles with a  hydrophilic shell made of intermittent maleic anhydride and octadecene co-polymer with subsequent coating with diglycidyl polyethylene glycol ether allowed for reduction of non-specific reaction of nanoparticles with plasma proteins. In its turn, it resulted in an increased time of their circulation in blood flow of small animals for up to 1 hour. With the Lewis lung carcinoma transfected to mice model we demonstrated аn in-life transportation of upconversion nanoparticles into the tumor with a  high degree of localization due to a passive EPR effect. The contrast of luminescent signal in the tumor compared to adjacent tissues was at least 70%. The possibility of visualization of upconverted nanoparticles up to 15 mm of biological tissue was shown. Conclusion: The optical imaging techniques with anti-Stokes photoluminescent markers ensure a  high contract real-time detection of tumor tissues that allows for their use for intra-operative diagnostics

Submicron polyacrolein particles in situ embedded with upconversion nanoparticles for bioassay

We report a new surface modification approach of upconversion nanoparticles (UCNPs) structured as inorganic hosts NaYF₄ codoped with Yb³⁺ and Er³⁺ based on their encapsulation in a two-stage process of precipitation polymerization of acrolein under alkaline conditions in the presence of UCNPs. The use of tetramethylammonium hydroxide both as an initiator of acrolein polymerization and as an agent for UCNP hydrophilization made it possible to increase the polyacrolein yield up to 90%. This approach enabled the facile, lossless embedment of UCNPs into the polymer particles suitable for bioassay. These particles are readily dispersible in aqueous and physiological buffers, exhibiting excellent photoluminescence properties, chemical stability, and also allow the control of particle diameters. The feasibility of the as-produced photoluminescent polymer particles mean-sized 260 nm for in vivo optical whole-animal imaging was also demonstrated using a home-built epi-luminescence imaging system.9 page(s

Quantitative imaging of single upconversion nanoparticles in biological tissue.

The unique luminescent properties of new-generation synthetic nanomaterials, upconversion nanoparticles (UCNPs), enabled high-contrast optical biomedical imaging by suppressing the crowded background of biological tissue autofluorescence and evading high tissue absorption. This raised high expectations on the UCNP utilities for intracellular and deep tissue imaging, such as whole animal imaging. At the same time, the critical nonlinear dependence of the UCNP luminescence on the excitation intensity results in dramatic signal reduction at (∼1 cm) depth in biological tissue. Here, we report on the experimental and theoretical investigation of this trade-off aiming at the identification of optimal application niches of UCNPs e.g. biological liquids and subsurface tissue layers. As an example of such applications, we report on single UCNP imaging through a layer of hemolyzed blood. To extend this result towards in vivo applications, we quantified the optical properties of single UCNPs and theoretically analyzed the prospects of single-particle detectability in live scattering and absorbing bio-tissue using a human skin model. The model predicts that a single 70-nm UCNP would be detectable at skin depths up to 400 µm, unlike a hardly detectable single fluorescent (fluorescein) dye molecule. UCNP-assisted imaging in the ballistic regime thus allows for excellent applications niches, where high sensitivity is the key requirement