Biophotonics 2020 Abstracts

Area 1 - Biophotonics
Full Papers
Nr: 3
Title:

Tissue Spectroscopy and Optical Clearing: Recent Progress for Clinical Applications
Authors:

Luís Oliveira and Valery Tuchin
Abstract: Tissue spectroscopy is a technique with long tradition in biophotonics [1]. The estimation of the wavelength dependency for the optical properties of biological tissues can be made from experimental spectral measurements through inverse simulations that use the Monte Carlo or the Adding-doubling methods [2]. Several of these studies have been made for various tissues and provided valuable information [3]. By combining tissue spectroscopy with the optical clearing technique further valuable information can be obtained [4]. Recent studies, based on tissue spectroscopy measurements, have been made to evaluate the diffusion properties of optical clearing agents in tissues. These studies provided differentiated diffusion properties for the same agent in normal and pathological tissues caused by different mobile water content in those tissues [5]. Recent publications show that spectroscopic measurements during optical clearing can be used to calculate the kinetics for the optical properties of tissues or to monitor the creation of two new tissue windows for clinical practice located at ultraviolet wavelengths [4]. By evaluating the third mechanism of optical clearing – the dissociation of proteins with deep ultraviolet spectroscopy it was recently possible to discriminate between normal and pathological tissues. This work is a review of the recent progresses in tissue spectroscopy as a result of measurements made during optical clearing treatments. [1] S. Musa, Computational Optical Biomedical Spectroscopy and Imaging, CRC Press, Boca Raton, 2015. [2] V. V. Tuchin, Tissue optics – Light Scattering Methods and Instruments for Medical Diagnosis, 3rd ed., SPIE Press, Bellingham, 2015. [3] S.L. Jacques, Optical properties of biological tissues: a review, Phys. Med. Biol. 58(11), R37-R61, 2013. [4] L. Oliveira and V. V. Tuchin, The Optical Clearing Method – A New Tool for Clinical Practice and Biomedical Engineering, Springer Briefs in Physics, Springer Nature, Cham (Switzerland), 2019. [5] I. Carneiro, S. Carvalho, R. Henrique, L. Oliveira, and V. V. Tuchin, A robust ex vivo method to evaluate the diffusion properties of agents in biological tissues, J. Biophot. 12(4): e201800333, 2019.
Nr: 9
Title:

Nanoscale Inspection of Toxic Amyloids by Tip-Enhanced Raman Spectroscopy

Authors:

Paolo Matteini, Cristiano D'Andrea and Roberto Pini
Abstract: Plasmon-enhanced spectroscopies rely on the resonance effects between a laser excitation and free conduction electrons in noble metal nanostructures, permitting to enhance the optical responses of molecules in their close proximity. Surface-Enhanced Raman Spectroscopy (SERS) is an ultrasensitive analytical technique that couples the unique features of Raman Spectroscopy in providing a description of chemical composition and structure of molecules with a huge signal enhancement due to isolated or assembled silver or gold nanoparticles. As a result, identification of trace amounts of molecules, including those of biomedical interest, becomes feasible [1-5]. The intrinsic chemical specificity of Raman Spectroscopy and the high signal sensitivity of SERS can be combined with the nanoscale spatial resolution of scanning probe microscopy (SPM) giving rise to Tip-Enhanced Raman Spectroscopy (TERS). In TERS the huge electromagnetic field localized on the apex of a sharp metallized tip is exploited to achieve compositional and structural information from the surface of nanosized samples [6]. In this work we show how to benefit from TERS as a surface-sensitive tool with spatial resolution on the nanoscale to inspect the spatial organization and surface character of individual protein oligomers causing cellular dysfunction and neurotoxicity. Aggregation processes of proteins leading to the formation of amyloid oligomers are nowadays considered primarily responsible for promoting synaptic failure and neuronal death associated with neurological disorders such as Alzheimer’s and Parkinson’s diseases. Therefore, unravelling the relationship between structure and neurotoxicity of protein oligomers becomes of primary importance in understanding the pathogenesis of the disease as well as in developing novel diagnostic and therapeutic strategies toward the earliest and pre-symptomatic stages of the disease. Our TERS investigation provides strong evidence of the presence of characteristic chemostructural determinants in toxic oligomers, which sheds new light on the mechanism by which they cause cellular impairment. Specifically, we provide direct assignment of specific aminoacid residues that are exposed on the surface of toxic amyloids, while appear buried in nontoxic amyloid forms. These residues, thanks to their outward disposition, might represent structural factors driving the pathogenic behaviour, including affecting cell membrane integrity and specific signal pathways in neurodegenerative conditions. [1] Guerrini L et al. ACS Appl Mater Interfaces 7 9420 (2015) [2] Matteini P et al. Nanoscale 8 3374 (2015) [3] Banchelli M et al. ACS Appl Mater Interfaces 8 2628 (2016) [4] Matteini P et al ACS Nano 11 918 (2017) [5] Banchelli M et al. Sci Rep 8, 1033 (2018) [6] D’Andrea C. et al, Small, 1800890 (2018).
Nr: 10
Title:

Assessing Tissue Mechanical Properties with Optical Coherence Elastography and Bruillion Spectroscopy
Authors:

Yogeshwari Ambekar, Raksha Raghunathan, Jitao Zhang, Giuliano Scarcelli and Kirill V. Larin
Abstract: The alternation of biomechanical properties of many tissues could indicate onset and progression of diseases. For example, stiffening of crystalline lens play a crucial role in its visual function and development presbyopia. Also, mechanical cues have been found to play crucial roles during embryo development, where the morphological evolution are contributed by both force and mechanical properties of local tissue. Optical coherence elastography and Brillouin microscopy have shown promise in determining the elasticity of biological samples, separately. In this work, we combined these methods to (1) map the longitudinal modulus of the neural tube tissue of mouse embryo and (2) quantify the Young’s modulus of whole lens ex vivo. We found that optical coherence elastography and Brillouin microscopy provide complementary information to yield truly quantitative measurements of tissue mechanical properties. This information can provide clues of early embryonic development as well as develop novel therapeutic procedures to treat congenital and tissue degenerative diseases.
Nr: 11
Title:

Glycerol and Glucose Diffusion in the Gingival Tissue of a Human Tooth Measured by an Optical Method: In Vitro Studies
Authors:

A. A. Selifonov, L. Oliveira and V. V. Tuchin
Abstract: Optical methods for visualizing and diagnosing tissues of the human body, based on the interaction of the optical radiation with biological tissues, are increasingly used in various fields of medicine: oncology, surgery, dermatology, neonatology, ophthalmology, pediatrics, dentistry, cosmetology, etc. and thereby stimulate the need for more detailed study of the optical characteristics of biological tissues and various cellular structures. The measured diffuse reflectance spectra can provide important information about the main biophysical parameters of the biological tissue under study: scattering and absorption coefficients, as well as the concentrations of optically active endogenous chromophores. However, significant scattering and partial absorption of radiation by biological tissues, including tissues of the gum mucosa and dentin of a human tooth, limit the transport of probe radiation to a sufficient depth of the tissue under study. To reduce the scattering of biological tissues, the optical clearing method is successfully used, which ensures an increase in the efficiency of optical methods in diagnostics and therapy. For immersion optical clearing of biological tissues, both hyperosmotic agents (glucose, sorbitol, glycerol, polyethylene glycol, propylene glycol, dimethyl sulfoxide) and isosmotic solutions (such as X-ray contrast agent iohexol, etc.) are used. Despite numerous studies related to the management of the optical properties of biological tissues, the problem of determining the diffusion coefficients of immersion fluids in biological tissues is still not well understood. Due to the complex multicomponent structure of biological tissues and the nonlinear nature of the processes of diffusion of immersion liquids, the determination of quantitative characteristics is a difficult task. In this work, we determined the effective diffusion coefficient of 99.5% glycerol and 40% glucose in the tissue of the human gingival mucosa in vitro, which amounted to (1.40±0.21)·10–6 cm2/s and (3.2±0.7)·10–6 cm2/s, using diffuse reflection spectroscopy and a free diffusion model in the wavelength range of 200 - 800 nm. It was found that the efficiency of optical clearing of the gum mucosa tissue under the influence of optical clearing agents has a maximum in the ultraviolet: when exposed to glycerol it was 2000% in the range 200-300 nm and for glucose, it was 300% in the range 300-400 nm.
Nr: 13
Title:

Twisted Structured Light for Tissue Diagnosis
Authors:

Igor Meglinski, Nicolas Vera, Juan P. Staforelli and Alex Doronin
Abstract: In turbid tissue-like scattering medium the conventional polarized light, scattered multiple number of times, is depolarized, and the depolarization rate depends strongly on the size and shape of scattering particles, as well as on the number of scattering events. In fact, the structure of light can be more complicated when the polarization of light across the laser beam can be radially or azimuthally polarized and carry orbital angular momentum (OAM). When these structured light beams, such as cylindrical vector beam (CVB) and/or Laguerre-Gaussian (LG) beams, propagates through a turbid tissue-like scattering medium, either anisotropic or inhomogeneous, the spin or angular momentum are changed that leads to spin-orbit interaction. The spin-orbit interaction leads to the mutual influence of the polarization and the trajectory of the light propagation. We investigate the applicability of using CVB and LG beams for characterization of biological tissues and their structural malformations associated with various dangerous diseases, including cancer. In current presentation propagation of CVB and LG beams in anisotropic turbid tissue-like scattering media is considered in comparison to conventional Gaussian beams. We demonstrate that contrast of visibility becomes at least twice higher by applying CVB and LG beams in comparison to the conventional tissue polarimetry approach utilizing Gaussian beams. We also show that when the twisted LG laser light propagates through normal and cancerous tissues the OAM is preserved with the noticeably different phase shift. Both experimental and theoretical results suggest that there is a high potential in application of structured light beams in tissue diagnosis. Nevertheless, the potential of twisted light for medical applications is far from being fully explored, in particular as the exact conditions and parameters for reliably recognizing disease onset are yet to be investigated in most cases.
Nr: 16
Title:

Optical Elastography: Imaging the Micro-mechanical Properties of Tissue
Authors:

Brendan Kennedy
Abstract: This talk will focus on the technical development of optical coherence elastography (OCE) and its application to both cancer imaging and mechanobiology. OCE utilises optical coherence tomography to map the deformation in tissue resulting from the application of a mechanical load. Use of a mechanical model allows us to generate 3D maps of tissue elasticity with a spatial resolution of 25-50 micrometres. As the mechanical properties of tissue are invariably modified on this scale, this emerging imaging capability has potential both as a novel diagnostic tool and as a means to provide insights on the genesis and progression of disease.
Nr: 19
Title:

Guiding Neurosurgeons During Deep Brain Stimulation Surgery with Optical Spectroscopy
Authors:

Daniel C. Cote and Damon DePaoli
Abstract: Differentiating tissue types is an important aspect of guiding medical interventions whether it be for disease diagnosis or for surgical guidance. However, diseased and healthy tissues are often hard to discriminate by human vision alone and surgical navigation can be difficult to accomplish in large organs where the target structure lies deep within the body. New methods that can increase certainty in such medical interventions are therefore of great interest to healthcare professionals. Optical spectroscopy is a tool which can be exploited to probe discriminatory information in tissue by analyzing light-tissue interactions with a spectral range, resolution and sensitivity much greater than the human eye. I will explain how I have leveraged optical spectroscopy to create, and improve, an optical guidance system for deep brain stimulation neurosurgery, specifically for the treatment of Parkinson’s disease. I will begin by describing how spectroscopic information can provide real-time feedback to a surgeon during the procedure, in the hopes of ultimately improving treatment outcome. To this end, I will present the investigation of two different spectroscopic modalities for optical guidance: diffuse reflectance spectroscopy, and coherent anti-Stokes Raman scattering spectroscopy. The advantages and disadvantages of both techniques will be discussed along with their promising translatability for this application. Experimental data in animals in primates will be shown.
Nr: 21
Title:

New Perspectives on Optical Imaging at Depth
Authors:

Kishan Dholakia
Abstract: We have seen great advances in photonics based methods for imaging and manipulation. However whislt improvements have been seen in beating the diffraction limit and gaining wide field imaging, imaging through scattering media remains a challenge. I will describe approaches that tackle this issue. In particular I will describe using a geometry where the excitation beam is orthogonal to the detection arm of the optical system has come to the fore. This method is known as light sheet imaging or selective plane illumination microscopy. This talk will describe the basic premises of this field and highlight how we may increase penetration using propagation invariant Bessel and Airy beams as well as multiphoton imaging, including particularly three photon excitation to obtain information at depth. In the second part of my talk I will move to an epi-fluorescent geometry and describe the use of temporal focusing with single pixel detection. This scheme which we term TRAFIX can image through turbid media without prior knowledge of the scattering properties present and can be implemented on both two and three photon imaging.
Nr: 23
Title:

Laser-Optics Methods for in Vitro and in Vivo Characterization of Microrheological Alterations of Blood Components in Microcirculatory System under Cardiovascular Diseases and Diabetes Mellitus
Authors:

Andrei E. Lugovtsov, Yury I. Gurfinkel and Alexander V. Priezzhev
Abstract: Nowadays the number of people suffering from diabetes mellitus and cardiovascular diseases (arterial hypertension, coronary heart disease) increases rapidly mainly due to unhealthy nutrition and lifestyle. Diabetes mellitus (DM) is a metabolic disease characterized by high blood sugar levels over a prolonged period. Arterial hypertension (AH), also known as high blood pressure, is a long-term medical condition in which the blood pressure in the arteries is persistently elevated. These diseases can lead to severe alterations of vitally important systems of the human organism including the cardiovascular system and resulting in damage to blood vessels and capillaries, impairment of blood hemorheology and microcirculation. Enhanced aggregation of erythrocytes and platelets is one of key factors, which determines the blood flow and thereby affects the blood rheology. The ability of erythrocytes to deform in shear flow conditions is second major property that affect blood microcirculation. Alterations in these properties lead to changing the blood viscosity and, as a consequence, to changes in capillary blood flow. This can lead to significant impairment of blood function, which increases a risk of occurrence of vascular concomitant diseases, and even the mortality. In this work, complex laser-optics studies of the factors determining the capillary blood flow in patients suffering from AH and DM were conducted. Laser aggregometry and diffractometry techniques were used to conduct in vitro measurements of aggregation and deformability characteristics of the erythrocytes on ensembles of cells [1, 2]. Double-channeled optical tweezers were used for in vitro measuring the aggregation speed as well as interaction forces during erythrocyte doublet formation on cellular level [3]. To visualize and quantitatively evaluate the capillary blood flow in vivo non-invasive capillaroscopy measurements in the nailfold vessels were conducted. It was shown that in AH and DM patients, the ability of erythrocytes to deform is slightly reduced while the aggregation speed and forces of the cells interaction are significantly increased relative to the control group. The blood microcirculation in nailfold capillaries is impaired as well. We shown that the alterations of the parameters measured in vivo and in vitro for patients with different stages of these diseases are interrelated. Good agreement between the results obtained with different techniques, and their applicability for the diagnostics of abnormalities of rheological properties of blood were demonstrated. Supported by the grant of the Russian Scientific Foundation # 18-15-00422. [1] A. Lugovtsov, Y. Gurfinkel, P. Ermolinskiy, A. Maslyanitsina, L. Dyachuk, and A. Priezzhev, Optical assessment of alterations of microrheologic and microcirculation parameters in cardiovascular diseases, Biomed. Opt. Express 10, 3974-3986 (2019). [2] A.V. Priezzhev, N.N. Firsov, J. Lademann, Light backscattering diagnostics of RBC aggregation in whole blood samples, Chapter 11 in Handbook of Optical Biomedical Diagnostics, Editor V. Tuchin, Washington: SPIE Press, 651 – 674 (2012). [3] K. Lee, M. Kinnunen, M.D. Khokhlova, E.V. Lyubin, A.V. Priezzhev, I. Meglinski, A. Fedyanin, Optical tweezers study of red blood cell aggregation and disaggregation in plasma and protein solutions, Journal of Biomedical Optics, 21(3), 035001 (2016).
Nr: 25
Title:

Laser-optic Methods For Characterization of Blood Microrheologic Properties
Authors:

A. Priezzhev, A. Lugovtsov, S. Nikitin, A. Semenov, P. Ermolinskiy and S. Shin
Abstract: Microrheologic parameters of blood essentially determine its microcirculation and, ultimately, the state of a person. Determining the microrheologic status of a person can and should become an integral part of his/her medical examination, especially in the presence of such socially significant diseases as cardiovascular, oncological, endocrinological (diabetes), etc. In this regard, the development and testing of new effective methods is an important task. The purpose of the work is to demonstrate the possibilities of using different laser-optic techniques to measure microrheologic parameters of blood related to the structure and dynamics of red blood cells (RBCs). In particular, we used diffuse light scattering (DLS), laser diffractometry (LD) and laser tweezers (LT) to measure parameters characterizing the aggregation of RBCs in blood samples obtained from healthy donors or patients suffering arterial hypertension, heart failure and/or diabetes mellitus. We measured the forces of aggregation and disaggregation of RBCs using LT [1,2] as well as the index of aggregation, the characteristic time of aggregation and the hydrodynamic strength of aggregates in whole blood using DLS. By means of LD we measured the RBC shear deformability [3]. Our results allow us to conclude that the laser-optic methods are an effective tool for studying the intrinsic properties of RBCs and their interactions, as well as for monitoring the pathological changes of their microrheologic parameters in vitro. This work was supported by the RFBR grant No. 19-52-51015. 1. Lee K, Danilina A.V., Priezzhev A.V. et al. Probing the red blood cells aggregating force with optical tweezers // IEEE Journal of Selected Topics in Quantum Electronics – 2016. – V.22, N3. – P. 7000106. 2. Lee K., Wagner Ch., Priezzhev A.V. Assessment of the “cross-bridge”-induced interaction of red blood cells by optical trapping combined with microfluidics // Journal of Biomedical Optics – 2017. – V.22, N9. – P. 091516. 3. Nikitin S.Yu., Ustinov V.D., Yurchuk Yu.S. New diffractometric equations and data processing algorithm for laser ektacytometry of red blood cells // Journal of Quantitative Spectroscopy & Radiative Transfer – 2016. – V.178. - P. 315–324.
Nr: 26
Title:

Diagnostic Value of Tissue Depolarization and Birefringence Obtained with Mueller Polarimetry
Authors:

Hee R. Lee, Meredith Kupinski, Angelo Pierangelo, Razvigor Ossikovski and Tatiana Novikova
Abstract: We explore the potential of Mueller imaging polarimetry operating in a visible wavelength range to become a new promising tool for tissue optical diagnostics. In particular, we focus our studies on different algorithms of polarimetric data post-processing and discuss the diagnostic value of both polarization (scalar retardance) and depolarization (total depolarization and eigenvalues of coherence matrix) parameters for accurate assessment of tissue conditions. We show that the detection performance of Mueller imaging polarimetry can be significantly increased by an appropriate choice of optimal optical markers.
Nr: 27
Title:

Ultra-Fine Needle Endoscope for Deep Interstitial Examination by Conventional, Autofluorescence Imaging and Raman Spectroscopy
Authors:

Alexandre Douplik
Abstract: This abstract is related to endoscopy, particularly needle endoscopy including biopsy imaging guidance, diagnostics and treatment monitoring. The key element is the acupuncture size hollow needle facility of 195-micron outer diameter size or smaller, which contains or deliver a light guiding 100-micron diameter fiber ultra-fine endoscope for conventional white color image (W) with autofluorescence (A) and Raman (R) diagnostic modalities (WAR) inside the needle for cancer or other types of pathologies’ diagnostics. From needle application studies it was noticed that smaller diameter needles cause less tissue destruction, pain and bleeding. From acupuncture practice we can be assumed that some damages/openings created by needles less than 200 microns of the outer diameter (34G and smaller) are self-reparable or non-critical for access from the surface and can be inserted up to 4-5 cm deep without excessive pain and bleeding at relatively fast healing. A conditional title for this technology is a Mosquito Bite Needle Endoscope (MBNE). MBNE uses a needle from a medical grade stainless steel of 45 mm or longer for deep tissue interrogation to obtain optical information from deep body structures at high resolution. Imaging by optical means is one of the diagnostic options when examination can be facilitated without ionization radiation at high resolution (ca. 5-10 microns). Such a high resolution can be critical if searching for small lesions and early pathological stages. The main limitation of optical imaging is light penetration depth into biological tissue: it is currently limited to 1-3 mm depending on the color of light used (wavelength range). This limit is prescribed by tissue optical properties, namely by high light scattering due to cell membranes, subcellular components, and interfaces between different tissue types, as well as high light absorption by intrinsic chromophores such as melanin and hemoglobin. Hence, most applications of optical imaging are limited to superficial or shallow examination of epithelium and mucosa, and comprise either handheld, free space, or microscopy like devices, or endoscopy. The MBNE technology proposes a deep interrogation of organ tissues at high resolution at a cost of minor invasive, practically painless and bleeding less, self-repairable examination. This approach is very novel and unique, supported by the cutting-edge technology of fine long length hollow needles engineering. In the case of diagnostic breast, thyroid or prostate imaging, the lesions in question are at depths from the skin surface of up to 3-5 cm. Although the recent development of other non-ionizing methods such as optoacoustic tomography promises to increase the interrogation depth to 1-2 cm, this method requires placement of the biological object into a water solution acting as a sound coupling media. Applications for this method in clinical settings of cancer surgery for breast, thyroid and prostate are therefore limited. Examples of preliminary autofluorescence and Raman results of interstitial needle endoscopy are provided. The MBNE facility allows a small invasive examination at the time of consultation, early diagnostics, reducing number of biopsies and surgery.
Nr: 28
Title:

Assessing Corneal Biomechanical Response to Simulated Ocular Pulsation
Authors:

Achuth Nair, Manmohan Singh, Salavat Aglyamov and Kirill V. Larin
Abstract: Introduction Understanding the biomechanical properties of the cornea can be useful for detecting ocular disease and monitoring therapeutic intervention. Mechanical analysis can be performed using elastography, a technique in which tissue deformation is quantified using an imaging modality. Ultrasound elastography and optical coherence elastography (OCE) have been previously utilized to assess the mechanical deformation of the cornea, typically in response to an external force. However, there is growing interest in assessing the tissue mechanical response to natural physiological forces. One such force is the corneal pulse that causes a distinct displacement in the tissue based on IOP fluctuations occuring at hearbeat rhythm. Corneal elasticity has been measured by inflation testing, but this method cannot assess dynamic changes in tissue stiffness. Time-reversal based passive elastography has been used in ocular tissues to assess stiffness in response to physiological forces but does not provide quantitative measurements of elasticity. Recently, ultrasound elastography has been used in response to simulated intraocular pressure fluctuations, but this method is limited by the resolution of its parent imaging modality. In this work, OCE is used to measure the corneal response to spatially varying fluctuations in IOP that simulates the heartbeat induced corneal pulse in the ex vivo porcine cornea. Materials and Methods Measurements were performed on an ex vivo porcine eye globe within 48 hours of enucleation from the animal. The eye globe was placed into a custom eye holder and cannulated with two syringes for IOP control. IOP was sinusoidally pulsed with an average pressure of 10 mmHg using a closed loop control system consisting of a syringe pump and a pressure transducer. A SD-OCT system with 6 µm axial resolution, 50 kHz line rate, and ~2 nm displacement sensitivity was used to detect the small displacements in the cornea induced by IOP fluctuation. OCT images were acquired using BM-mode imaging at 50 kHz frame rate during IOP pulsation. Motion within the cornea was detected using the complex OCT signal, and the axial displacement was calculated based on the complex phase difference between an initial reference frame and each consecutive frame. Axial strain was calculated from displacement using a linear regression-based method. Results and Discussion Our results indicate a cyclical corneal biomechanical response that corresponds to mechanical forces induced by fluid infusion and withdrawal into the ocular cavity due to the simulated IOP fluctuation. The measured displacement was translated to strain. Average displacement within the cornea ranged from -1.6 ± 0.5 µm to 0.9 ± 0.3 µm, suggesting that there is an overall compression at the anterior of the cornea and relaxation at the posterior of the cornea over the entire pulse. While we have yet to quantify corneal mechanical properties, our results suggests a mechanical gradient through the cornea. Conclusion In this work, we show preliminary data for analyzing the biomechanical properties of the cornea during dynamic intraocular pressure changes that simulate the corneal pulse. Our work suggests that corneal strain varies axially during pulsatile IOP fluctuation similar to the ocular pulse. Future work will consider changes in mechanical properties at different baseline IOPs and will include quantitative metrics to assess elasticity. The results shown here suggests that corneal biomechanics can be measured in vivo without the need for external excitation.
Nr: 29
Title:

Imaging Device for Skin Subsurface Analysis based on Spatial Angular Filter (SAF)
Authors:

Alexandre Douplik, Irina Schelkanova and Aditya Pandya
Abstract: Lens-based imaging technologies are currently used in various applications although the use of a lens at high resolutions reduces the depth of field, so only a shallow plane can be imaged, and this image can be easily distorted by motion. An example of such an application is a human skin examination where imaging of dermal structures becomes particularly difficult due to a turbid environment that highly scatters the light, reducing contrast and blurring the image. The contemporary methods resolving these issues are either relatively expensive such as Optical Coherence Tomography (OCT) or Photoacoustic Tomography (PAT) or insufficient to take a high resolution, high contrast images at required depth of focus. Our team developed a lensless imaging device capable of microscopic resolution with a greater depth of field of view. This solution involved development of an optomechanical approach capable of imaging light propagation in a medium that strongly scatters electromagnetic waves. This approach does not require complex image processing methods and can therefore be used with any type of a camera or light source. Our invention is focused on recovering an absorption contrast within highly scattering media via gating imaging pixels and illumination delivery which was named by us as “spatial angle filtering” (SAF) technology. Through gating of photons exiting the scattering media using a restricted numerical aperture (NA) fiber optic plate (FOP), using dilutions of Intralipid (1-4 v/v%) and a USAF resolution target, we have shown that by reducing the NA of the FOP from 0.55 to 0.17, the interrogation depth improves ca. 2 times using trans-illumination. As the scattering increased (to 3-4% of Intralipid concentration, which is comparable with the scattering in human skin), this ratio decreased prompting a non-linear dependence of the interrogation depth to the scattering. Both experimental and simulation results are presented. The primer field of application that drove creation and development of this technology is biomedical optics. This technology can be exploited either stand alone or complementary to Optical Coherence Tomography (OCT) or Fluorescence Imaging for biomedical applications. Apart of biomedical applications we presume that this technology can be applied for a deep interrogation to significantly enhance the vision ability via and inside highly scattering media such as fog, cloud, muddy water, scattering solid matters like paper, various biological tissue etc. In general, SAF does not require active optoelectronics, or a complex image processing and can be described as a passive optical device that can be added on to various camera and spectral systems. It is a cost-effective solution that is particularly effective for improving interrogation depth in turbid media and can also be extended to applications in multiple industries. The SAF device does not require focusing and therefore does not have any strict dimensional requirements, making it feasible both for miniature applications such as endoscopy and large-scale applications such as industrial imaging.
Nr: 30
Title:

Improving Optoacoustic Imaging Performance with Deep Learning
Authors:

N. Davoudi, X. L. Deán-Ben and D. Razansky
Abstract: The rapidly evolving field of optoacoustic imaging and tomography is driven by a constant need for better imaging performance in terms of resolution, speed, sensitivity, depth and contrast. In practice, data acquisition strategies commonly involve sub-optimal sampling of the tomographic data, resulting in inevitable performance trade-offs and diminished image quality. We developed a new framework for efficient recovery of image quality from sparse optoacoustic data based on a deep convolutional neural network and demonstrate its performance with whole body in vivo mouse imaging. To generate accurate high-resolution reference images for optimal training, a full-view tomographic scanner capable of attaining superior cross-sectional image quality from living mice was devised. When provided with reconstructed images from substantially undersampled data and limited-view scans, the trained network was capable of enhancing the visibility of arbitrarily oriented structures and restoring the expected image quality. Notably, the network also eliminated some reconstruction artefacts present in the densely sampled signal reference images that were reconstructed. No comparable gains were achieved when the training was performed with synthetic or phantom data, underlining the importance of training with high-quality in vivo images acquired by full-view scanners. The new method can benefit numerous optoacoustic imaging applications by mitigating common image artefacts, enhancing anatomical contrast and image quantification capacities, accelerating data acquisition and image reconstruction approaches, while also facilitating the development of practical and affordable imaging systems. The suggested approach operates solely on image-domain data and thus can be seamlessly applied to artefactual images reconstructed with other modalities.
Nr: 32
Title:

Shedding Light on Radiotherapy: Optical Coherence Angiography (OCA) to Assess Tissue Functional Response to Radiation
Authors:

Alex Vitkin
Abstract: Despite widespread use and tremendous technological and radiobiological advances, radiation therapy as a cancer treatment remains somewhat ‘blind’ – its relative success and outcome are often not known for weeks or months following treatment. Can advanced analysis of in-vivo optical coherence tomography (OCT) images of irradiated animal tumours detect early functional changes induced by radiotherapy? If yes, can these be used to ‘shed light on radiotherapy’ and optimize / personalize its treatment delivery? We address these questions by analyzing OCT temporal and spatial signal statistics (for microvascular and microstructural imaging, respectively). In this talk we will demonstrate the imaging and analysis pipeline, show representative preclinical and clinical results, and discuss implications.

Area 1 - Biophotonics

Full Papers
Paper Nr: 2
Title:

Optimization of a Cold Atmospheric Plasma Treatment to Selectively Affect the Viability of Skin Cancer Cells
Authors:

Sara Pereira, Paulo A. Ribeiro and Susana Sério
Abstract: Cold atmospheric plasmas (CAPs) are a specific type of non-thermal plasmas mainly composed by reactive oxygen and nitrogen species (ROS and RNS, respectively), UV radiation and charged particles. In the last years, liquids treated by CAPs (indirect CAPs treatments) have attracted a significant interest in oncology due to its ability to kill cancer cells with an effectiveness similar to direct irradiation of the cells by cold plasmas. It is important to point out that indirect treatments have the advantage of avoiding the effects of UV radiation and the electrical fields present in plasmas being their effects mainly dependent on the ROS and RNS produced in the liquid phase. To better understand the mechanisms behind the interaction between CAPs, treated liquids and cells, it was engineered a plasma jet device and studied the vulnerability of different cell lines to the culture medium previously exposed to CAPs. For that, it was analysed the concentration of H2O2 produced during the treatments by means of calorimetric assays and evaluated the influence of using different working parameters such as volume of medium and gap. According to the obtained results it could be observed that the cancer cell line (Met-1) in study is more sensitive to the liquids treated by CAPs than the non-cancer one (HGF-1), which in the particular case of this jet, seems to be mainly related with the concentration of RNS species produced in the liquids during the plasma exposure since the concentration of the H2O2 produced is very low.
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Nr: 24
Title:

Optical Radiomic Signatures Derived from OCT Images to Improve Identification of Melanoma
Authors:

Peter E. Andersen
Abstract: Malignant melanoma is by far the most dangerous type of skin cancer. Currently, the gold standard to diagnose melanoma in the clinic is excisional biopsy and histopathologic analysis. Approximately 15-30 benign lesions are biopsied to diagnose each melanoma. Additionally, biopsies are invasive and result in pain, anxiety, scarring and disfigurement of patients, and they can be a financial burden to the health care system. Among several imaging techniques developed to enhance melanoma diagnosis, optical coherence tomography (OCT) with its high-resolution and intermediate penetration depth can potentially provide required diagnostic information, noninvasively. We propose an image analysis algorithm, ‘optical properties extraction (OPE)’ that drastically improves the specificity and sensitivity of OCT by identifying unique optical radiomic signatures pertinent to melanoma detection. We evaluate the performance of the algorithm using several tissue-mimicking phantoms. We then test the OPE algorithm with sixty-nine human subjects and demonstrate that melanoma can be differentiated from benign nevi with 97% sensitivity, and 98% specificity.
Nr: 1
Title:

Investigation of Reproductive Process and Early Development in Vivo using Functional Optical Coherence Tomography
Authors:

Irina V. Larina
Abstract: Optical imaging provides unique opportunities to investigate dynamics of reproduction and early development with single cell resolution without application of exogenous agents in animal models. Ovulation, fertilization, and pre-implantation pregnancy are fundamental reproductive processes of clinical importance. While research has shed light on the cellular and molecular mechanisms mediating these events, any conclusions regarding dynamics of mammalian fertilization, gamete/embryo transfer, which takes place deep inside the body, are extrapolated and do not necessarily represent the native state. Toward this end, using functional optical coherence tomography (OCT), we recently established a set of unique methods for in vivo imaging of the female mouse reproductive tract. Our approach allows for live, dynamic volumetric imaging of the mouse Fallopian tube (oviduct) with micro-scale spatial resolution. Recently, we established a tomographic imaging technique capable of mapping both the cilia location and the cilia beat frequency in the intact mouse oviduct in vivo. We utilize both the amplitude and the frequency position of the major peak from the ciliary motion spectrum, and reconstruct, for the first time, high-resolution mapping of both cilia location and CBF through tissue layers in vivo. We established a method of sperm activity measurement, which relies on analysis of unique tortious sperm trajectories. We introduced standard deviation (SD) of the direction variation (SDofDV) as a measure of activity. This method was used in vivo to directly track multiple motile sperm inside the mouse oviduct, revealing new biological findings. Potentially this study will provide new insight on the process of mammalian fertilization in its native state and lead to a better understanding of pathologies resulting in infertility.
Nr: 4
Title:

Deep Stain-Free Cancer-Cell Grading
Authors:

Natan T. Shaked
Abstract: We used clinical-ready digital holographic setups to acquire live healthy and cancer cells of different grades without staining. We then designed new machine learning classifiers that can work with small training sets for rapidly retrieving the cell grade from its stain-free quantitative phase map. The method has a great potential to aid cancer diagnosis in imaging flow cytometry.
Nr: 5
Title:

Overcoming the Abbe Diffraction Limit in Terahertz Imaging of Soft Tissues
Authors:

K. I. Zaytsev, N. V. Chernomyrdin, G. M. Katyba, I. N. Dolganova, V. N. Kurlov and V. V. Tuchin
Abstract: Dimensions of the structural elements of tissues (i.e. organels, separate cells and their agglomerates, microfibrils etc.) are usually small at the scale posed by the terahertz (THz) wavelengths [1,2]. Such tissue elements could not be resolved using conventional modalities of THz imaging, which rely on the diffraction-limited lens- and mirror-based optical systems [3]. This pushes further developments into the realm of THz imaging of tissues with strongly sub-wavelength spatial resolution. In this talk, we review our developments in the high-resolution THz imaging of tissues: – solid immersion microscopy [4–8]; – imaging based on the photonic jet effect [9–11]; – scanning-probe near-field microscopy based on flexible sapphire fibers [11–14]. We compare these novel THz imaging modalities considering their spatial resolution, energy efficiency, and applicability in biophotonics. Finally, we present the results of sub-wavelength resolution THz imaging of soft biological tissues, including benign and malignant neoplasms with different localization and nosology. [1] Progress in Quantum Electronics 62, 1 (2018). [2] Journal of Optics (2019, in press), DOI: 10.1088/2040-8986/ab4dc3. [3] Review of Scientific Instruments 88, 014703 (2017). [4] Applied Physics Letters 110, 221109 (2017). [5] Applied Physics Letters 113, 111102 (2018). [6] Optics & Spectroscopy 126, 560 (2019). [7] N.V. Chernomyrdin et al., “Numerical analysis and experimental study of terahertz solid immersion microscopy,” Optical Engineering (2019, under review). [8] Proceedings of SPIE 11164, 111640H (2019). [9] Optical Materials Express 7, 1820 (2017). [10] APL Photonics 2, 056106 (2017). [11] Applied Physics Letters 114, 031105 (2019). [12] Proceedings of SPIE 11088, 110880I (2019). [13] Proceedings of SPIE 11164, 111640G (2019). [14] Proceedings of SPIE 11088, 110880I (2019).
Nr: 6
Title:

Four-dimensional Optical Coherence Tomography and Optogenetic Control of Cardiodynamics in Mouse Embryos
Authors:

Andrew L. Lopez and Irina V. Larina
Abstract: Congenital heart defects (CHD) are common and occur in nearly 1% of all live births. Moreover, cardiovascular (CV) failures are the leading cause of birth defect-related deaths in infants. It is established that biomechanical stimuli are critical regulators of CV development. Thus, defining how mechanical factors are integrated with genetic pathways to coordinate mammalian heart tube development and function will factor into strategies for new therapeutic interventions to treat/prevent CHD in humans. To address this critical need, we have established optical approaches for live, high-resolution imaging and manipulation of mouse embryo cardiodynamics. Our approach combines live mouse embryo culture with structural and functional OCT imaging, second harmonic generation (SHG) microscopy and optogenetics. We have developed methods for acquisition, synchronization, of the developing beating heart with spatial resolution of about 4μm, and 4D angiography approach to visualize dynamics of blood flows without staining within beating embryonic hearts. Toward understanding of the pumping mechanism, one of intriguing findings revealed that the heart wall at neighboring sites accelerates in coordination to generate very high retrograde flows, which was only found at the site of valve development, suggesting that it might be critical in this process. The extracellular matrix (ECM), and particularly fibrillar collagen, are central to heart biomechanics, regulating tissue strength, elasticity and contractility. Our studies using SHG microscopy revealed cardiac fibers, such as collagen, coinciding with areas of higher mechanical load through the earliest stages of heart development, and suggested regulatory role of heart contraction in establishment of mechanical homeostasis. To control cardiac contraction, we have established optogenetic cardiac pacing in embryos that express the light-activated, transmembrane channel—Channelrhodopsin2—only in the heart. We have integrated a pulsed 473nm laser with our lab-built optical coherence tomography system to control embryonic heart beat frequency, and generate 4D (3D+time) images of the heart to extract structural and functional information such as heart wall and blood flow velocity to validate and measure changes in heart biomechanics. Paced hearts will be imaged with SHG and image processed to determine changes in collagen content and organization due to altered contraction. This work will be the first-time use of a non-contact approach to test the consequences of altered mechanics in early heart development.
Nr: 7
Title:

Super-resolved and Extended Depth of Focus Label-free Imaging in Microscopy, Microendoscopy and Photoacoustics
Authors:

Zeev Zalevsky
Abstract: Imaging systems as well as human vision system have limited capability for separation of spatial features and this information can also be extracted only from depth limited range. The reasons to the resolution and depth of focus limitations are related to the effect of diffraction i.e. the finite dimensions of the imaging optics as well as the geometry of the sensor. In my talk I will present novel photonic approaches and means to exceed the above-mentioned resolution and show how those concepts can be adapted to microendoscopy as well as to microscopy related configurations. In the case of microendoscopy, I will show how projection of wavelength and time dependent codes can enhance the resolution of the collected light. This concept of super resolved imaging will also be demonstrated for imaging through biological scattering medium such as biological tissues and liquids as blood. The projected wavelength and time dependent high-resolution encoding patterns are sent via laser-based illumination fiber while the collected light is collected via ultra-thin multicore imaging fiber-based endoscope. In the case of microscopy I will present how the resolution limit can go much below sub-wavelength bound towards nanoscopic imaging while using label-free configurations involving time multiplexing (time dependent light collection) based upon label-free non-static nano-particles either moving in uncontrolled Brownian motion or being manipulated with light (optical tweezers). In the case of photoacoustic I will show how the above-mentioned super resolved concepts can be combined with interference based extension of depth of focus and be applied for photo acoustic imaging configurations.
Nr: 8
Title:

Fluorescence Lifetime Techniques in Clinical Interventions
Authors:

Laura Marcu
Abstract: This presentation overviews fluorescence lifetime spectroscopy and imaging techniques for label-free in vivo characterization of biological tissues. Emphasis is placed on recently developed devices and methods enabling real-time characterization and diagnosis of diseased tissues during clinical interventions. I will present studies conducted in human patients demonstrating the ability of these techniques to provide a rapid in-situ evaluation of tissue biochemistry and their potential to guide surgical and intravascular procedures. Current results demonstrate that intrinsic fluorescence can provide useful contrast for intraoperative delineation of brain tumors head and neck tumors and breast cancer. Finally, I will present results from the first-in-human study that shows the potential of a multispectral fluorescence lifetime method for image-guided augmented reality in trans-oral robotic surgery (TORS).
Nr: 12
Title:

Applications of Master Slave Optical Coherence Tomography
Authors:

Adrian Podoleanu, Adrian Bradu, Ramona Cernat, Manuel Marques and Sylvain Rivet
Abstract: Master Slave (MS) optical coherence tomography (OCT) was introduced to deliver en-face OCT images direct, not possible using A-scan based conventional OCT technology. The MS method relies on comparison of channeled spectra, where a processor is allocated to each depth of interest in the object to be imaged, equipped with a mask and a comparator. The mask represents the channeled spectrum obtained with a two paths interferometer, each path terminated on a mirror, for a specific optical path difference. The comparators in each processor deliver signal strengths proportional to the similarity of acquired channeled spectrum from the object with each mask. An immediate advantage is that no resampling - linearization is needed in case of the channeled spectrum is chirped. Examples will be given using both spectrometer based and swept source OCT systems. In addition, tolerance to dispersion in the interferometer will also be demonstrated. This property, together with that of tolerance to chirp allows MS-OCT more consistence in achieving theoretical axial resolution when employing broadband optical sources such as supercontinuum lasers, than the conventional spectral domain OCT method based on Fourier transformations. Because en-face images are generated in real time, the volume of the object can be assembled from stacks of such images. This capability also allows reviving the OCT/SLO (scanning laser ophthalmoscopy) concept, a landmark of time domain OCT. In this case, both OCT and SLO channels process signals employing spectral domain principles, so they exhibit superior sensitivity in comparison with the original OCT/SLO reports. For ultra high speed or long coherence length swept source OCT, that would require an expensive, multi – GS/s digitiser, MS-downconversion OCT will be presented. This allows real time en-face delivery using a much lower cost and lower sampling rate digitiser, at the level of the sweeping rate.