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Objectives: The paper focuses on the development of technology of adipose tissue optical clearing using different complex hyperosmotic optical clearing agents and tissue permeability enhancers., Methods: To quantify optical clearing efficiency, reduced scattering coefficient was estimated from the ex vivo spatially resolved backreflectance measurements using a multi-distant fiber optical device. Tissue morphology modification was monitored with the help of histological studies., Results: Kinetics and efficiency of the optical clearing were evaluated for ex vivo abdominal fat tissue at action of dimethyl sulfoxide, diatrizoic acid, metrizoic acid, sucrose, and fructose solutions accompanied by tissue permeability enhancers, such as various modes of fractional laser microablation and sonophoresis and their combinations. Histological studies allowed us to find agents that cause minimal morphological changes of the adipose tissue., Conclusions: Maximal optical clearing efficiency of 83.5% was observed for the samples subjected to action of the fructose-ethanol solution during 90 min in combination with gentle modes of fractional laser microablation and sonophoresis., (© 2024 Wiley Periodicals LLC.)

Photoacoustic flow cytometry is one of the most effective approaches to detect "alien" objects in the bloodstream, including circulating tumor cells, blood clots, parasites, and emboli. However, the possibility of detecting high-amplitude signals from these objects against the background of blood depends on the parameters of the laser pulse. So, the dependencies of photoacoustic signals amplitude and number on laser pulse energy (5-150 μJ), pulse length (1, 2, 5 ns), and pulse repetition rate (2, 5, 10 kHz) for the melanoma cells were investigated. First, the PA responses of a melanoma cell suspension in vitro were measured to directly assess the efficiency of converting laser light into an acoustic signal. After it, the same dependence with the developed murine model based on constant rate melanoma cell injection into the animal blood flow was tested. Both in vivo and in vitro experiments show that signal generation efficiency increases with laser pulse energy above 15 μJ. Shorter pulses, especially 1 ns, provide more efficient signal generation as well as higher pulse rates. A higher pulse rate also provides more efficient signal generation, but also leads to overheating of the skin. The results show the limits where the photoacoustic flow cytometry system can be effectively used for the detection of circulating tumor cells in undiluted blood both for in vitro experiments and for in vivo murine models., (© 2023 International Society for Advancement of Cytometry.)

Objectives: The development of compact diagnostic probes and instruments with an ability to direct access to organs and tissues and integration of these instruments into surgical workflows is an important task of modern physics and medicine. The need for such tools is essential for surgical oncology, where intraoperative visualization and demarcation of tumor margins define further prognosis and survival of patients. In this paper, the possible solution for this intraoperative imaging problem is proposed and its feasibility to detect tumorous tissue is studied experimentally., Methods: For this aim, the sapphire scalpel was developed and fabricated using the edge-defined film-fed growth technique aided by mechanical grinding, polishing, and chemical sharpening of the cutting edge. It possesses optical transparency, mechanical strength, chemical inertness, and thermal resistance alongside the presence of the as-grown hollow capillary channels in its volume for accommodating optical fibers. The rounding of the cutting edge exceeds the same for metal scalpels and can be as small as 110 nm. Thanks to these features, sapphire scalpel combines tissue dissection with light delivering and optical diagnosis. The feasibility for the tumor margin detection was studied, including both gelatin-based tissue phantoms and ex vivo freshly excised specimens of the basal cell carcinoma from humans and the glioma model 101.8 from rats. These tumors are commonly diagnosed either non-invasively or intraoperatively using different modalities of fluorescence spectroscopy and imaging, which makes them ideal candidates for our feasibility test. For this purpose, fiber-based spectroscopic measurements of the backscattered laser radiation and the fluorescence signals were carried out in the visible range., Results: Experimental studies show the feasibility of the proposed sapphire scalpel to provide a 2-mm-resolution of the tumor margins' detection, along with an ability to distinguish the tumor invasion region, which results from analysis of the backscattered optical fields and the endogenous or exogenous fluorescence data., Conclusions: Our findings justified a strong potential of the sapphire scalpel for surgical oncology. However, further research and engineering efforts are required to optimize the sapphire scalpel geometry and the optical diagnosis protocols to meet the requirements of oncosurgery, including diagnosis and resection of neoplasms with different localizations and nosologies., (© 2021 Wiley Periodicals LLC.)

Background: The analysis of recent studies on plasmonic photothermal therapy (PPT) after intravenous administration of gold nanorods (GNRs) has demonstrated that the effectiveness of nanoparticle-assisted laser hyperthermia depends on a correct dosage strategy of nanoparticle administration. Accumulation of GNRs in tumor tissue dramatically increases the local heating of the tumor without damage to healthy tissues. However, the optimal doses of GNR intravenous injections (IVIs) for effective accumulation in tumors, and optimal protocols of PPT are not designed yet. The current study aims to improve the efficacy of PPT in tumor-bearing rats using multiple fractional intravenous administration of GNRs., Materials and Methods: For PPT experiments, the GNRs with aspect ratio of 4.1 were functionalized with thiolated polyethylene glycol (PEG) and their suspensions were used for multiple fractional intravenous administration in outbred albino male rats with experimental model of rat liver cancer (cholangiocarcinoma line PC-1). Doppler ultrasonography was performed to characterize the vascularity of transplanted rat tumors before any treatment. After a final injection of GNRs, tumor was irradiated during 15 minutes by 808-nm NIR diode laser at a power density 2.3 W/cm 2 . The animals were withdrawn from the experiment and sampling of tissues for morphological study and gold accumulation was performed 24 hours and 3 weeks after PPT., Results: The multiple IVIs of gold nanorods and further PPT of transplanted cholangiocarcinoma provided significant damage to tumor tissue resulting in pronounced necrotic mass and retardation of the tumor growth. More importantly, the proposed PPT protocol had low toxicity as evidenced by histological examination of internal organs. The efficiency of PPT depends on the presence of newly formed vasculature as revealed by the Doppler ultrasound investigation., Conclusion: The repeatable IVIs promote greater of GNR accumulation within the tumor thus resulting in higher PPT efficacy. Accompanying ultrasonography can be useful for prognosis and monitoring of treatment. Lasers Surg. Med. 50:1025-1033, 2018. © 2018 Wiley Periodicals, Inc., (© 2018 Wiley Periodicals, Inc.)

Background and Objectives: Laser radiation is often used to provide micro and nanoparticle delivery into the skin for medical and cosmetic purposes. This technique inherently has limited speed and effective penetration. We proposed and investigated a new method of rapid delivery of solid microparticles, nanoparticles and liquids into tissue through multiple microchannels created by a fractional laser microablation (FLMA) using Er:YAG-laser. The dependence of microchannel depth on laser pulse temporal structure and number of pulses and dermal coloration changes are studied in this paper., Study Design/materials and Methods: Microchannels created in the porcine skin in vitro by a fractional Er:YAG-laser were used to deliver Zirconium oxide (ZrO2) microparticles or hydrocortisone solution. Each laser pulse consisted of subpulses. Number of laser pulses (Np) and subpulses (Nsp) can be adjusted. The enhancement of delivery is expected due to hydrodynamic impact of laser pulse on the layer of the aqueous suspension of the particles or hydrocortisone solution placed on the skin surface. For color investigation, we used standard CIE Lab parameter analysis., Results: The relationship between microchannel depth in the skin and number of laser pulses and subpulses was established. We found that free filling of microchannels with ZrO2-particle suspension has a low speed of ∼4 × 10(-5)  mm/s. Particle delivery into microchannels induced by the hydrodynamic shock waves generated by Er:YAG-laser pulses is carried out with a high speed of 28.5 mm/s. We also found that skin color at ZrO2 -particle delivery differs from color of the intact skin, namely: the parameter L, which characterizes the "lightness" increased by 9 ± 1%; parameter a, which characterizes the "redness" decreased by 38 ± 4%; and parameter b, which characterizes the "yellowness" decreased by 21 ± 2%. The effective delivery of hydrocortisone was demonstrated using fluorescence method technique., Conclusion: Multi-beam laser-induced hydrodynamic shock waves generated by Er:YAG-laser pulses on the layer of the aqueous suspension of the particles or solution of a high molecular weight drug placed on the skin can be used for their rapid delivery into the skin., (© 2015 Wiley Periodicals, Inc.)

Flow cytometry (FCM) has been a fundamental tool of biological discovery for many years. Invasive extraction of cells from a living organism, however, may lead to changes in cell properties and prevents studying cells in their native environment. These problems can be overcome by use of in vivo FCM, which provides detection and imaging of circulating normal and abnormal cells directly in blood or lymph flow. The goal of this review is to provide a brief history, features, and challenges of this new generation of FCM methods and instruments. Spectrum of possibilities of in vivo FCM in biological science (e.g., cell metabolism, immune function, or apoptosis) and medical fields (e.g., cancer, infection, and cardiovascular disorder) including integrated photoacoustic-photothermal theranostics of circulating abnormal cells are discussed with focus on recent advances of this new platform., (Copyright © 2011 International Society for Advancement of Cytometry.)

Background and Objectives: Tissue optical clearing technique based on immersion of tissues into optical clearing agents (OCAs) can reduce the scattering and enhance the penetration of light in tissue. However, the barrier function of epidermis limits the penetration of OCAs, and hence is responsible for the poor optical clearing efficacy of skin by topical action. In this study, a variety of light irradiation was applied to increase permeability of agents in skin and improve the optical clearing efficacy., Study Design/materials and Methods: Different light sources with different dose, i.e, CO(2) laser, Nd:YAG laser (532 and 1,064 nm) with different pulse modes and Intense Pulsed Light (IPL) (400-700 and 560-950 nm) were used to irradiate rat skin in vivo, and then glycerol was applied onto the irradiated zone. VIS-NIR spectrometer was utilized to monitor the changes of reflectance. In vitro skin samples were also irradiated by Q-switched Nd:YAG laser (1,064 nm) and then treated by glycerol for 10-60 minutes. Based on the measurement of the reflectance and transmittance of the samples, the optical properties of skin and penetration depth of light were calculated., Results: Results show that photo-irradiation with appropriate dose combining with the following glycerol treatment is able to reduce in vivo skin reflectance. Compared with the control group, the maximal changes in reflectance are ninefold at 575 nm and eightfold at 615 nm, respectively, which were caused by Q-switched 1,064-nm Nd:YAG laser irradiation and following glycerol treatment. The results for in vitro skin demonstrate that the joint action can significantly increase the optical penetration depth in samples., Conclusions: The combination of Q-switched Nd:YAG (1,064 nm) laser and glycerol could enhance optical skin clearing efficacy significantly. This study provides a non-invasive way to improve the optical clearing of skin, which will benefit the skin optical therapy.

Compared with blood tests, cell assessment in lymphatics is not well-established. The goal of this work was to develop in vivo lymph tests using the principles of flow cytometry. Cells in living animals were counted by laser (420-2,300 nm) generation of photoacoustic (PA) signals in individual cells hydrodynamically focused by lymph valves into a single file flow, and using endogenous absorption as intrinsic cell-specific markers, or gold nanorods, nanoshells, and carbon nanotubes as multicolor probes. PA data were verified by high-speed transmission, photothermal, and fluorescent imaging. Counting of melanoma and immune-related cells in normal, apoptotic, and necrotic states in lymphatics in vivo was demonstrated to have the unprecedented sensitivity as one metastatic cell among millions of white blood cells. The time-resolved PA spectral identification of flowing cells was achieved using multicolor labels and laser pulses of different wavelengths and time delays. Multiparameter, noninvasive, portable flow cytometer can be used for preclinical studies on animals with the potential of translation to humans for in vivo PA mapping of colorless lymph vessels and sentinel nodes with simultaneous single cell detection and metastasis assessment without labeling or use of contrast dyes and/or novel low-toxic multicolor probes with different absorption spectra., (Copyright 2008 International Society for Advancement of Cytometry.)

Background: Photothermal (PT) cytometry has recently demonstrated great potential for the label-free detection of nonfluorescent cells under static conditions. The goal of our investigation was to expand this technique to the detection of flowing cells in vitro., Methods: Cells in flow were irradiated with short, tunable laser pulses (420-2,300 nm, 8 ns), and the absorbed energy was detected by monitoring of the temperature-dependent variations in the refractive index in the cells with a second, collinear probe beam in two modes: (a) PT imaging of single cells with a pulsed probe beam (639 nm, 13 ns) and (b) thermolens monitoring of the integral PT responses from individual cells as whole with a continuous-wave probe beam (633 nm, 2 mW)., Results: PT flow cytometry at the current speed of analysis of 10 cell/s, with the capability to image selected cells of interest flowing at velocities up to 2 m/s, demonstrated the capability for (a) label-free detection of flowing single cells (e.g., blood and cancer cells) on the basis of the differences in their endogenous absorption properties, (b) identification of cells labeled with gold nanoparticles, (c) rapid cell viability testing, (d) aggregation immunoassay, and (e) optimization of selective nanophotothermolysis., Conclusions: PT cytometry can be extended to the study of cells in flow. This new technique increases the speed of cell analysis approximately 10(2) times over that of conventional PT technique, with the potential to achieve a rate of 10(4)-10(5) cells/s in specific PT applications, which has previously been realized only with cells under static conditions., ((c) 2007 International Society for Analytical Cytology.)

Background and Objectives: Strong light scattering in skin prevents precise targeting of optical energy in therapeutic and diagnostic applications. Optical immersion based on matching refractive index of scattering centers with that of surrounding matter through introduction of an exogenous index-matching agent can alleviate the problem. However, slow diffusion of the index-matching agent through skin barrier makes practical implementation of this approach difficult. We propose a method of accelerating penetration of the index-matching compounds by enhancing skin permeability through creating a lattice of micro-zones (islets) of limited thermal damage in the stratum corneum (SC)., Study Design/materials and Methods: A flash lamp (intense pulsed light) system and an island mask with a pattern of absorbing centers (center size approximately 75-120 microm, lattice pitch approximately 450-500 microm) were used to create the lattice of islets of damage (LID). Index-matching agents, such as glucose solution, propylene glycol solution, and glycerol solution, were applied., Results: Experimental results of optical clearing ex vivo rat and pig skin, and ex vivo and in vivo human skin are presented. Optical transmission spectra of the skin samples with LID were measured during some 2 hours after application of index-matching chemical agents. In order to assess and compare the clearing rate under different treatment and clearing agents we calculated the quantity that we call "relative transmittance": T(rel) = I(t)(lambda)/I(0)(lambda), were I(t)(lambda) is the intensity measured at elapsed time t. The dynamics of relative transmittance of skin samples at 470 and 650 nm shows that the implementation of limited thermal damage technique leads to a 3-10-fold increase of optical clearing (rise of transmittance) rate compared to the results obtained when the samples were treated with high-intensity light pulses but without the use of island damage mask (IDM). It was observed from the plotted spectra of relative transmittance that the maximum increase of transmitted light intensity has been obtained with glucose solution as a clearing agent. Noteworthy is the difference in the trend of spectral curves: relative transmittance spectrum for glycerol reveals, on the whole, a greater slope which may be indicative of higher extent of index matching between the scattering centers and base material for this index-matching agent. Under the transillumination of the skin sample by the wide flat beam the more effective clearing (the increase of transmitted intensity) is attained within the hemoglobin absorption bands; with the narrow quasi-collimated beam the higher relative transmittance was observed over the intervals of minimum absorption., Conclusions: The use of specially designed island mask combined with non-laser intensive pulse irradiation produces a lattice of islands of limited thermal damage in SC that substantially enhances the penetration rate of topically applied index-matching agents. The suggested technique gave comparable magnitudes of clearing dynamics enhancement for glucose solution, glycerol solution, and propylene glycol solution applied to mammalian skin.

Flow cytometry is a well-established, powerful technique for studying cells in artificial flow in vitro. This review covers a new potential application of this technique for studying normal and abnormal cells in their native condition in blood or lymph flow in vivo. Specifically, the capabilities of the label-free photothermal (PT) technique for detecting and imaging cells in the microvessel network of rat mesentery are analyzed from the point of view of overcoming the problems of flow cytometry in vivo. These problems include, among others, the influences of light scattering and absorption in vessel walls and surrounding tissues, instability of cell velocity, and cells numbers and positions in a vessel's cross-section. The potential applications of this new approach in cell biochemistry and medicine are discussed, including molecular imaging; studying the metabolism and pathogenesis of many diseases at a cellular level; and monitoring and quantifying metastatic and apoptotic cells, and/or their responses to therapeutic interventions (e.g., drug or radiation), in natural biological environments., ((c) 2006 Wiley-Liss, Inc.)

Background and Objectives: Objective evaluation of the pigmentation index (PI) and the eythema index (EI) of human skin is a prerequisite for successful optimization of laser- and intense-pulsed-light (IPL)-based treatment modalities in dermatology., Study Design/materials and Methods: We describe a three-wavelength technique for determining PI and EI as well as its particular implementation using LEDs operating at wavelengths of 560, 650, and 710 nm and a large-area photodiode. The instrument has been evaluated both in vitro and in vivo., Results: In vitro, good correlation between the measured indices and results obtained with commercially available techniques has been observed. In addition, linearity of the PI with melanin concentration in the phantom medium up to 7 x 10(-3) nm(-1) (defined as a slope of the optical density spectrum) has been established. In vivo, feasibility of using the technique for predicting the minimal erythema dose (MED), minimal phototoxic dose (MPD), and the threshold of epidermal damage in a photothermal treatment has been demonstrated., Conclusions: The data suggest that the technique has a substantial potential as a method of pre-treatment diagnostics for photochemical and photothermal procedures., (Copyright 2004 Wiley-Liss, Inc.)

Background and Objectives: The near-infrared (NIR) laser radiation due to its high penetration depth is widely used in phototherapy. In application to skin appendages, a high selectivity of laser treatment is needed to prevent light action on surrounding tissues. Indocyanine green (ICG) dye may provide a high selectivity of treatment due to effective ICG uploading by a target and its narrow band of considerable absorption just at the wavelength of the NIR diode laser. The goal of this study is to demonstrate the efficacy of the NIR diode laser phototherapy in combination with topical application of ICG suggested for soft and thermal treatment of acne vulgaris., Study Design/materials and Methods: Twenty-two volunteers with facile or back-located acne were enrolled. Skin sites of subjects were stained by ICG and irradiated by NIR laser-diode light (803 or 809 nm). One mg/ml solution of ICG was applied for 5 or 15 minutes to the cleaned skin site. Untreated, only stained and only light irradiated skin areas served as controls. For soft acne treatment, the low-intensity (803 nm, 10-50 mW/cm(2), 5-10 minutes) or the medium-intensity (809 nm, 150-190 mW/cm(2), 15 minutes) protocols were used. The single and multiple (up to 8-9) treatments were provided. The individual acne lesions were photothermally treated at 18 W/cm(2) (803 nm, 0.5 seconds) without skin surface cooling or at 200 W/cm(2) (809 nm, 0.5 seconds) with cooling., Results: The observations during 1-2 months showed that soft acne treatment decreased the number of active elements, reduced erythema and inflammation, and considerably improved the skin state without any side effects. At high power densities (up to 200 W/cm(2)), ICG stained acne inflammatory elements were destroyed for light exposures of 0.5 seconds., Conclusions: Based on the concept that hair follicle, especially sebaceous gland, can be intensively and selectively stained by ICG due to dye diffusion through pilosebaceous canal and its fast uptake by living microorganisms, by vital keratinocytes of epithelium of the canal and sebaceous duct, and by rapidly proliferating sebocytes, new technologies of soft and thermal acne lesions treatment that could be used in clinical treatment of acne were proposed., (Copyright 2003 Wiley-Liss, Inc.)