Methods for optical skin clearing in molecular optical imaging in dermatology

This short review describes recent progress in using optical clearing (OC) technique in skin studies. Optical clear-ing is an efficient tool for enhancing the probing depth and data quality in multiphoton microscopy and Raman spec-troscopy. Here, we discuss the main mechanisms of OC, its safety, advantages, and limitations. The data on the OC effect on the skin water content are presented. It was demonstrated that 70% glycerol and 100% OmnipaqueTM 300 reduce the water content in the skin. Both OC agents (OCAs) significantly affect the strongly bound and weakly bound water. However, OmnipaqueTM 300 causes considerably less skin dehydration than glycerol. In addition, the results of examination of the OC effect on autofluorescence in two-photon excitation and background fluorescence in Raman scattering at different skin depths are presented. It is shown that OmnipaqueTM 300 is a promising OCA due to its ability to reduce background fluo-rescence in the upper skin layers. The possibility of multimodal imaging combining optical methods and OC technique is discussed

Combined laser speckle imaging and fluorescent intravital microscopy for monitoring acute vascular permeability reaction

Optical clearing agents (OCAs) and many chemicals are widely used in functional diagnosis of skin tissues. Numerous studies are associated with the transcutaneous diffusion of OCA in epidermal, dermal, and hypodermal tissues, which results in changing their optical properties. In addition, an objective approach that is suitable for screening the influence of utilized OCA, as well as various chemical agents, synthetics, and nanomaterials, on blood and lymph flows is highly desirable. In our study, a highly sensitive laser speckle imaging (LSI) system and fluorescent intravital microscopy (FIM) were used team-wise to inspect the acute skin vascular permeability reaction in mouse ear during the local application of OCA on the skin surface. Fluorescent contrast material administrated intravenously was used for quantitatively assessing the intensity of vascular permeability reaction and the strength of skin irritation. The obtained results suggest that a combined use of LSI and FIM is highly effective for monitoring the cutaneous vascular permeability reaction, with great potential for assessment of allergic reactions of skin in response to interactions with chemical substances

Time-space Fourier κω' filter for motion artifacts compensation during transcranial fluorescence brain imaging

Intravital imaging of brain vasculature through the intact cranium in vivo is based on the evolution of the fluorescence intensity and provides an ability to characterize various physiological processes in the natural context of cellular resolution. The involuntary motions of the examined subjects often limit in vivo non-invasive functional optical imaging. Conventional imaging diagnostic modalities encounter serious difficulties in correction of artificial motions, associated with fast high dynamics of the intensity values in the collected image sequences, when a common reference cannot be provided. In the current report, we introduce an alternative solution based on a time-space Fourier transform method so-called K-Omega. We demonstrate that the proposed approach is effective for image stabilization of fast dynamic image sequences and can be used autonomously without supervision and assignation of a reference image

A Robust Method for Adjustment of Laser Speckle Contrast Imaging during Transcranial Mouse Brain Visualization

Laser speckle imaging (LSI) is a well-known and useful approach for the non-invasive visualization of flows and microcirculation localized in turbid scattering media, including biological tissues (such as brain vasculature, skin capillaries etc.). Despite an extensive use of LSI for brain imaging, the LSI technique has several critical limitations. One of them is associated with inability to resolve a functionality of vessels. This limitation also leads to the systematic error in the quantitative interpretation of values of speckle contrast obtained for different vessel types, such as sagittal sinus, arteries, and veins. Here, utilizing a combined use of LSI and fluorescent intravital microscopy (FIM), we present a simple and robust method to overcome the limitations mentioned above for the LSI approach. The proposed technique provides more relevant, abundant, and valuable information regarding perfusion rate ration between different types of vessels that makes this method highly useful for in vivo brain surgical operations

Evaluation of handwriting peculiarities utilizing laser speckle contrast imaging

Functional handwriting is a process involving various complex interactions between physical, cognitive and sensory systems. Since muscular motion is of a peculiar nature for each person, handwriting properties, such as pencil pressure and speed of writing, can be considered as a unique marker of identity. Moreover, impairments of handwriting in many cases are connected to neurodevelopmental disorders such as attention deficit hyperactivity disorder, developmental coordination disorder, autism spectrum disorders, Parkinson's disease, etc. From this point of view, investigations of handwriting kinematics and pressure can be highly important for both forensic science and medicine. Commonly, the kinematic and pressure features of handwriting are evaluated using a graphics tablet with a stylus or electronic pens. The production of such devices is quite expensive. Therefore, the development of new methods for individual handwriting analysis is an important and current goal. Laser speckle contrast imaging (LSCI) is a powerful method, which is sensitive to both motion and pressure. Since the developed technique requires the use of only a simple laser diode and camera for image acquisition, LSCI is a cost-effective and practical tool for handwriting analysis. In the current letter we present a robust LSCI-based method for handwriting pressure and kinematics evaluation. The introduced approach was validated by an Archimedean spiral writing task

Advances in Dynamic Light Scattering Imaging of Blood Flow

Dynamic light scattering (DLS) is a well known experimental approach uniquely suited for the characterization of small particles undergoing Brownian motion in randomly inhomogeneous turbid scattering medium, including water suspension, polymers in solutions, cells cultures, and so on. DLS is based on the illuminating of turbid medium with a coherent laser light and further analyzes the intensity fluctuations caused by the motion of the scattering particles. The DLS-based spin-off derivative techniques, such laser Doppler flowmetry (LDF), diffusing wave spectroscopy (DWS), laser speckle contrast imaging (LSCI), and Doppler optical coherence tomography (DOCT), are exploited widely for non-invasive imaging of blood flow in brain, skin, muscles, and other biological tissues. The recent advancements in the DLS-based imaging technologies in frame of their application for brain blood flow monitoring, skin perfusion measurements, and non-invasive blood micro-circulation characterization are overviewed. The fundamentals, breakthrough potential, and practical findings revealed by DLS-based blood flow imaging studies, including the limitations and challenges of the approach such as movement artifacts, non-ergodicity, and overcoming high scattering properties of studied medium, are also discussed. It is concluded that continued research and further technological advancements in DLS-based imaging will pave the way for new exciting developments and insights into blood flow diagnostic imaging

Beyond life: Exploring hemodynamic patterns in postmortem mice brains

We utilize Laser Speckle Contrast Imaging (LSCI) for visualizing cerebral blood flow in mice during and post-cardiac arrest. Analyzing LSCI images, we noted temporal blood flow variations across the brain surface for hours postmortem. Fast Fourier Transform (FFT) analysis depicted blood flow and microcirculation decay post-death. Continuous Wavelet Transform (CWT) identified potential cerebral hemodynamic synchronization patterns. Additionally, non-negative matrix factorization (NMF) with four components segmented LSCI images, revealing structural subcomponent alterations over time. This integrated approach of LSCI, FFT, CWT, and NMF offers a comprehensive tool for studying cerebral blood flow dynamics, metaphorically capturing the ‘end of the tunnel’ experience. Results showed primary postmortem hemodynamic activity in the olfactory bulbs, followed by blood microflow relocations between somatosensory and visual cortical regions via the superior sagittal sinus. This method opens new avenues for exploring these phenomena, potentially linking neuroscientific insights with mysteries surrounding consciousness and perception at life's end

Speckle dynamics under ergodicity breaking

Laser speckle contrast imaging (LSCI) is a well-known and versatile approach for the non-invasive visualization of flows and microcirculation localized in turbid scattering media, including biological tissues. In most conventional implementations of LSCI the ergodic regime is typically assumed valid. However, most composite turbid scattering media, especially biological tissues, are non-ergodic, containing a mixture of dynamic and static centers of light scattering. In the current study, we examined the speckle contrast in different dynamic conditions with the aim of assessing limitations in the quantitative interpretation of speckle contrast images. Based on a simple phenomenological approach, we introduced a coefficient of speckle dynamics to quantitatively assess the ratio of the dynamic part of a scattering medium to the static one. The introduced coefficient allows one to distinguish real changes in motion from the mere appearance of static components in the field of view. As examples of systems with static/dynamic transitions, thawing and heating of Intralipid samples were studied by the LSCI approach

Ex-vivo confocal Raman microspectroscopy of porcine skin with 633/785-NM laser excitation and optical clearing with glycerol/water/DMSO solution

Confocal Raman microspectroscopy (CRM) with 633- and 785-nm excitation wavelengths combined with optical clearing (OC) technique was used for ex-vivo study of porcine skin in the Raman ¯ngerprint region. The optical clearing has been performed on the skin samples by applying a mixture of glycerol and distilled water and a mixture of glycerol, distilled water and chemical penetration enhancer dimethyl sulfoxide (DMSO) during 30 min and 60 min of treatment. It was shown that the combined use of the optical clearing technique and CRM at 633nm allowed one to preserve the high probing depth, signal-to-noise ratio and spectral resolution simultaneously. Comparing the e®ect of di®erent optical clearing agents on porcine skin showed that an optical clearing agent containing chemical penetration enhancer provides higher optical clearing e±ciency. Also, an increase in treatment time allows to improve the optical clearing e±ciency of both optical clearing agents. As a result of optical clearing, the detection of the amide-III spectral region indicating well-distinguishable structural di®erences between the type-I and type-IV collagens has been improved

Stokes-correlometry analysis of biological tissues with polycrystalline structure

Utilizing Stokes-correlometry analysis a new diagnostic approach has been introduced for quantitative assessment of polarization images of histological sections of optically anisotropic biological tissues with different morphological structures and physiological conditions. The developed approach is based on the quantitative assessment of coordinate and phase distributions of the Stokes vector of scattered light. A combined use of statistic, correlation, and fractal analysis is used for resolving variations in optical anisotropy of biological samples. The proposed combined application of the statistical, correlation, and fractal-based evaluates of spatial distributions of `single-point' polarization azimuth, ellipticity, and `two-point' Stokes vector parameters of polarization images of biological tissues histological sections demonstrates a high accuracy (Ac ≥ 90%) in monitoring of optical anisotropy variations within biological tissues