This PDF file contains the front matter associated with SPIE Proceedings Volume 10506, including the Title Page, Copyright information, Table of Contents, and Conference Committee listing.

Breast cancer has become a major cause of death among women. The lifetime risk of a woman developing this disease has been established as one in eight. The most useful way to reduce breast cancer death is to treat the disease as early as possible. The existing methods of early diagnostics of breast cancer are mainly based on screening mammography or Magnetic Resonance Imaging (MRI) periodically conducted at medical facilities. In this paper the authors proposing a new approach for simple breast cancer detection. It is based on skin stimulation by sound waves, illuminating it by laser beam and tracking the reflected secondary speckle patterns. As first approach, plastic balls of different sizes were placed under the skin of chicken breast and detected by the proposed method.

We developed the concept of temporal depth imaging and defined non-flat signals as signals with different dispersion values as a function of time. We demonstrated how shifting the timing of a time lens makes it possible to retrieve the dispersion value of each point in the signal, which is equivalent to a 3D imaging system. Finally, we demonstrated how a time lens array can retrieve these values with a single measurement by comparing the different images obtained by the time lens array.

We developed temporal super-resolution technique by adopting super-resolution techniques from space to time. Similar to spatial optics, where knowledge about the basic building blocks of the image can lead to better resolution, as demonstrated by localization microscopy techniques. We are utilizing our knowledge on the shape and duration of the pulses to retrieve a super-resolution image in the time domain of an input signal. The resolution of our time-lens is much lower than the needed resolution to obtain the signal but never-the-less we obtain a temporal image with high resolution.

Cell samples flowing along a microfluidic tube are scanned with an optical coherence tomography (OCT) system and their correlation times in M-mode scans are calibrated. In particular, the variations of correlation time with waiting time after 5 and 10 % ethanol are applied to the cell samples are compared for understanding the evolution of cell morphology in the cell death pathways of apoptosis and necrosis, respectively. Also, Au nanorings (NRIs) are taken up by cells for increasing the scattering strength in OCT scanning and hence increasing the signal-to-noise ratio. It is found that when cells are incubated with 5 % ethanol, the correlation time keeps decreasing with waiting time and then increases at 7 hours. On the other hand, when cells are incubated with 10 % ethanol, the correlation time keeps decreasing with waiting time all the way up to 7 hours. This difference may imply that the correlation time from OCT scanning may be determined not only by the size of cell fragment, but also by the smoothness of the cell fragment in a scale of several hundred nm during the apoptosis and necrosis processes. In particular, the results imply that the surface smoothness of the apoptotic bodies formed at the final stage of an apoptosis process is higher than that of the cell fragments formed at the final stage of a necrosis process. This OCT scanning technique has the potential application to the determination of cell death condition with the function similar to cell flow cytometry.

Far-field super-resolution imaging techniques such as stimulated emission depletion (STED), stochastic optical reconstruction microscopy (STORM), and photoactivation localization microscopy (PALM) etc., have enabled fluorescence labeled nanoscale imaging and analysis. However, it is necessary or preferable to develop label-free high resolution imaging techniques to avoid the problems of phototoxicity with live cells and moreover, for many non-biological applications where fluorescent labelling is simply not feasible. Recently, by capturing the exponentially decaying evanescent waves and propagating it to far field, novel imaging technique like hyperlens and microsphere contacting technologies, micro-fiber and nanowire illumination technology have brought new opportunities to label-free super resolution imaging, but still suffer from weak signal compared to background noise. On the other hand, there is lack of criteria reported to quantify the imaging quality of label free far-field nanoscopy, that will slow down the developing of label-free far-field nanoscopy. We introduce CNR first to label-free far-field nanoscopy to quantify the imaging performance and investigate the key influencing elements systematically. In our study, we investigated and analyzed the key elements to achieve high CNR label-free wide-field far-field sub-diffraction imaging. By optimizing the key factors including polarization, materials, as well as fabrication conditions, sub-diffraction imaging with resolution of 122 nm in a large FOV has been achieved experimentally. This work has provided an efficient and convenient way to realize high contrast wide-field far-field label-free super-resolution based on evanescent wave illumination.

In the era of Single-Cell Biology or Cytomics (as termed in the early 2000th) it is aimed to characterize every individual cell of an organism. Not only that, but for the Cell Atlas (or Human Cytome) Project it is intended to also characterize each cell type in various physiological states (quiescence, activation, aging, death). This ambitious goal is only reachable by combining the virtues of different scientific and technological disciplines such as high-end microscopy and cytometry, single cell genomics and proteomics, and bioinformatics. In the present discovery phase it becomes clear that the more cellular parameters (surface or intracellular markers such as peptides, sugar residues, lipids, gene expression profiles etc.) can be measured per individual cells the more functionally distinct cell subsets are identified. So far it’s a great bonanza for discovery as it is not foreseeable how many distinct cell subtypes exist in an organism but for unraveling it needs new technologies for highly multiplexed single cell analysis. The ideal technology for unraveling subsets in heterogeneous cell populations based on the complex combination (barcode) of a multitude of markers is flow and image cytometry. This field amazingly evolved in the last years. Here I will give an overview on the recent developments in complex instrumentation, dyes for specifically tagging cell properties (fluorochromes, nanoparticles, rare earth elements) and new bioinformatics approaches to visualize and analyze high-dimensional data. I will also emphasize on how different approaches could be combined for enabling the highest level of multiplexing.

Atherosclerosis, the leading cause of morbidity and mortality of cardiovascular disease, occur due to hardening and narrowing of arteries for development of vulnerable plaques made of cholesterols, tissue macrophages, foam cells and smooth muscle cells. Early detection of atherosclerosis is essential for proper treatment. Our group has already reported about the potential application of the non-invasive diffusion reflection (DR) technique in the detection of atherosclerosis using gold nanorods (GNRs) as contrast agent in carotid artery injured mice model. The basics of the study lie on the uptake GNRs by macrophages that located at the vulnerable plaques, which act as a good absorption contrast for DR measurement. Accumulations of GNRs cause changes in the optical property of the tissues and in turn cause changes in DR profile. In this study, we report the potential application of DR measurement in the detection of atherosclerosis in high-fat diet mice. Here, we have used PEG-coated GNRs with absorption maxima around 660nm. The time kinetics showed that after 24h of GNR injection the DR can find the atherosclerotic plaques and with time (up to 72h) the GNR accumulation in plaques were faded out, but still can be detectable by DR. Our result strongly suggests that in future DR can be used for early detection of atherosclerosis.

Light-activated theranostic constructs provide a multi-functional platform for optical imaging and phototherapeutic applications. Our group has engineered nano-sized vesicles derived from erythrocytes that encapsulate the FDAapproved near infrared (NIR) absorber indocyanine green (ICG). We refer to these constructs as NIR erythrocytemimicking transducers (NETs). Once photo-excited by NIR light these constructs can transduce the photons energy to emit fluorescence, generate heat, or induce chemical reactions. In this study, we investigated fluorescence imaging of NETs embedded within tumor phantoms using spatial frequency domain imaging (SFDI). Using SFDI, we were able to fluorescently image simulated tumors doped with different concentration of NETs. These preliminary results suggest that NETs can be used in conjunction with SFDI for potential tumor imaging applications.

Design of multifunctional nanoparticles for multimodal in-vivo imaging and advanced targeting to diseased single cells for massive parallel processing nanomedicine approaches requires careful overall design and a multilayered approach. Initial core materials can include non-toxic metals which not only serve as an x-ray contrast agent for CAT scan imaging, but can contain T1 or T2 contrast agents for MRI imaging. One choice is superparamagnetic iron oxide NPs which also allow for convenient magnetic manipulation during manufacturing but also for re-positioning inside the body and for single cell hyperthermia therapies. To permit real-time fluorescence-guided surgery, fluorescence molecules can be included. Advanced targeting can be achieved by attaching antibodies, peptides, aptamers, or other targeting molecules to the nanoparticle in a multilayered approach producing “programmable nanoparticles” whereby the “programming” means controlling a sequence of multi-step targeting methods. Addition of membrane permeating peptides can facilitate uptake by the cell. Addition of “stealth” molecules (e.g. PEG or chitosan) to the outer surfaces of the nanoparticles can permit greatly enhanced circulation times in-vivo which in turn lead to lower amounts of drug exposure to the patient which can reduce undesirable side effects. Nanoparticles with incomplete layers can be removed by affinity purification methods to minimize mistargeting events in-vivo. Nanoscale imaging of these manufactured, multifunctional nanoparticles can be achieved either directly through superresolution microscopy or indirectly through single nanoparticle zeta-sizing or x-ray correlation microscopy. Since these multifunctional nanoparticles are best analyzed by technologies permitting analysis in aqueous environments, superresolution microscopy is, in most cases, the preferred method.

Imaging the distribution of gold nanoparticles attached to target cells is of critical importance for numerous studies, including plasmonic cell transfection, drug delivery, and plasmonic photothermal and photoionization therapy. Thanks to their unique optical properties, gold nanoparticles can be characterized by a variety of optical spectroscopy techniques; the characterization of individual nanoparticles, however, requires high-resolution microscopy techniques such as dark-field microscopy, two-photon excitation fluorescence microscopy, or, alternatively, transmission electron microscopy for resolving the particles’ structure and shape. When attached to living cells, individual gold nanoparticles are much more difficult to resolve because their scattering properties are still low, even under plasmonic resonance illumination, compared to the characteristic scattering of cellular organelles. To overcome this problem, some researchers tend to increase nanoparticle concentration considerably; such practice often results in high cytotoxicity and considerable particle aggregation. The talk will outline the difficulties associated with gold nanoparticle detection in cells and propose solutions that integrate two-photon excitation fluorescence microscopy, absorption spectroscopy and scattering electron microscopy, for mapping particle distribution within the cells and estimating the level of particle aggregation. We will discuss our approach for estimating nanoparticle distribution within three-dimensional cell cultures, where light penetration is limited and electron microscopy is ineffective. Two-photon microscopy of these cultures after conjugation to gold nanoparticle revealed subsurface aggregates that were widely dispersed across the culture. These findings assist our current efforts to better control particle delivery into tissue phantoms for effective targeting of cancer cells by plasmonic phototherapy.

Precise polarimetric imaging of polarization-sensitive nanoparticles is essential for resolving their accurate spatial positions beyond the diffraction limit. However, conventional technologies currently suffer from beam deviation errors which cannot be corrected beyond the diffraction limit. To overcome this issue, we experimentally demonstrate a spatially stable nano-imaging system for polarization-sensitive nanoparticles. In this study, we show that by integrating a voltage-tunable imaging variable polarizer with optical microscopy, we are able to suppress beam deviation errors. We expect that this nano-imaging system should allow for acquisition of accurate positional and polarization information from individual nanoparticles in applications where real-time, high precision spatial information is required.

Carbon dots (CDs) are a new class of carbon nanomaterials and luminescent materials. Due to their fascinating properties, CDs can be widely applied in many fields, including bioimaging, biosensors, and photocatalysis. Performance characteristics of CDs are mainly concentrated in diluted solutions for a long time, it is possible to play advantages of small size effect and excellent cell permeability. However, the single morphology limits the development and applications cannot make progress further. To take full advantage of the low-toxicity, strong-luminescence CDs in the fields of biophotonics and optoelectronics, it is desirable to embed them in an appropriate solid matrix. Recently, due to its simplicity, versatility and cost-effectiveness, the electrospinning technique has been widely used to fabricate nanofibers from a wide variety of materials. To our knowledge, there are very few reports available on CDs embed in one-dimensional nanofibers. Herein, we use silane pre-functionalized CDs (SiCDs) and select silane as alkoxide precursor, then the SiCD/SiO2 composite nanofibers are prepared by combining sol-gel method with electrospinning technique. The resulting product may possess new physical or chemical properties from the two components. The fluorescence quantum yield of the SiCD/SiO2 composite nanofibers is as high as 90% and the fluorescence lifetime can reach up to about 6 ns. Bioapplication experiment confirm the low cytotoxicity, well bioimaging and biocompatibility of these nanofibers. It is expected that this one-dimensional composite fibers may have more wide applications in bioimaging, textile, and optical devices for their novel geometrical structure and luminescent properties.

Owing to its sensitivity and precise control at the nanoscale, polyelectrolytes have been immensely used to modify surfaces. Polyelectrolyte multilayers are generally water made and are easy to fabricate on any surface by the layer-by-layer (LbL) self-assembly process due to electrostatic interactions. Polyelectrolyte multilayers or PEMs can be assembled to form ultrathin membranes which can have potential applications in water filtration and desalination [1-3]. Hydration in PEMs is a consequence of both the bulk and surface phenomenon [4-7]. Bulk behavior of polymer membranes are well understood. Several techniques including reflectivity and contact angle measurements were used to measure the hydration in the bulk of polymer membranes [4, 8]. On the other hand their internal organization at the molecular level which can have a profound contribution in the transport mechanism, are not understood well. Previously, we engineered a technique, which we refer to as Bright-field Nanoscopy, which allows nanoscale optical imaging using local heterogeneities in a water-soluble germanium (Ge) thin film (~ 25 nm thick) deposited on gold [8]. We use this technique to study the water transport in PEMs. It is understood that the surface charge and outer layers of the PEMs play a significant role in water transport through polymers [9-11]. This well-known ‘odd-even’ effect arising on having different surface termination of the PEMs was optically observed with a spatial resolution unlike any other reported previously [12]. In this communication, we report that on increasing the etchant’s concentration, one can control the lateral etching of the Ge film. This allowed the visualization of the nanoscale internal organization in the PEMs. Knowledge of the internal structure would allow one to engineer polymer membranes specific to applications such as drug delivering capsules, ion transport membranes and barriers etc. We also demonstrate a mathematical model involving a surface permeability term which captures the experimentally observed odd-even effect.

Optical biosensing is a growing area of research known for its low limits of detection. Among optical sensing techniques, fluorescence detection is among the most established and prevalent. Fluorescence imaging is an optical biosensing modality that exploits the sensitivity of fluorescence in an easy-to-use process. Fluorescence imaging allows a user to place a sample on a sensor and use an imager, such as a camera, to collect the results. The image can then be processed to determine the presence of the analyte. Fluorescence imaging is appealing because it can be performed with as little as a light source, a camera and a data processor thus being ideal for nontrained personnel without any expensive equipment. Fluorescence imaging sensors generally employ an immunoassay procedure to selectively trap analytes such as antigens or antibodies. When the analyte is present, the sensor fluoresces thus transducing the chemical reaction into an optical signal capable of imaging. Enhancement of this fluorescence leads to an enhancement in the detection capabilities of the sensor. Diatoms are unicellular algae with a biosilica shell called a frustule. The frustule is porous with periodic nanopores making them biological photonic crystals. Additionally, the porous nature of the frustule allows for large surface area capable of multiple analyte binding sites. In this paper, we fabricate a diatom based ultra-sensitive fluorescence imaging biosensor capable of detecting the antibody mouse immunoglobulin down to a concentration of 1 nM. The measured signal has an enhancement of 6× when compared to sensors fabricated without diatoms.

Integrated Mach-Zehnder interferometers, ring resonators, Bragg reflectors or simple waveguides are commonly used as photonic biosensing elements. They can be used for label-free detection relating the changes in the optical signal in realtime, as optical power or spectral response, to the presence and even the quantity of a target analyte on the surface of the photonic waveguide. The label-free method has advantages in term of sample preparation but it is more sensitive to spurious effects such as temperature and refractive index sample variation, biological noise, etc. Label methods can be more robust, more sensitive and able to manipulate the biological targets.

In this work, we present an innovative labeled biosensing technique exploiting magnetic nano-beads for enhancement of sensitivity over integrated optic microrings. A sandwich binding is exploited to bring the magnetic labels close to the surface of the optical waveguide and interact with the optical evanescent field.

The proximity and the quantity of the magnetic nano-beads are seen as a shift in the resonance of the microring. Detection of antibodies permits to reach a high level of sensitivity, down to 8 pM with a high confidence level. The sizes of the nano-beads are 50 to 250 nm. Furthermore, time-varying magnetic fields permit to manipulate the beads and even induce specific signals on the detected light to easy the processing and provide a reliable identification of the presence of the desired analyte. Multiple analytes detection is also possible.

It is required to reduce the excitation volume of fluorescence with enhanced field intensity to apply fluorescence correlation spectroscopy (FCS) technique to the investigation of biological molecules. Field localization induced by plasmonic nanostructures enables measurements of molecular dynamics under high concentration exhibiting high signal-to-noise (SNR) ratio. To achieve this goal, we have investigated the feasibility of plasmonic monomer and dimer nanostructures for FCS techniques. We have studied field enhancement and localization induced by different gold monomer arrays whose shapes were circle, rhombus and triangle and also gold dimer arrays which had a gap of 10 nm. These plasmnoic nanostructures were considered to be on the gold film and glass substrate with chrome adhesion layer. We could shift the peak wavelength of field enhancement by changing the dimensions of nanostructures to spectrally overlap the field enhancement to excitation and emission spectrum of fluorophores. In the case of dimer configuration having a 90-nm diameter and a 20-nm height, we have induced the near-field localization with a light source at 671 nm whose dimension was 18×6×6 nm^3 with an enhanced field intensity by 500 times in comparison with a incident light. The field distribution was analyzed numerically and experimentally using finite-difference-time-domain method and near-field scanning optical microscope. We could measure the diffusion coefficients of 50-nm fluorescent beads with improved SNR which was found to be 44.6×10-14 m^2/s. Results would include the diffusion mapping of fluorescence molecules using imaging FCS technique to show the plasmnoic nanostructures is applicable to nanoscale FCS for study of molecular biology.

Supercritical angle fluorescence (SAF) is a near-field collection method that has surface sensitivities similar to or better than near-field excitation techniques like TIRF and waveguide based excitation. SAF is emitted by fluorophores that are a few hundred nanometers away from an interface, above the critical angle, into the higher index material. SAF decreases exponentially with increasing distance from the interface and is therefore more sensitive to molecules near the surface. Although a lot of research has used SAF for biosensing and microscopy, the angular dependence of SAF on both the surface and bulk fluorescence contributions hasn’t been experimentally studied. We present a method that measures the surface selectivity of SAF in the presence of bulk fluorophores. Two different fluorophores were used. One was bound to the surface and the other was suspended in the bulk. The spectrum was measured at discrete points in the back focal plane (BFP) and the contribution of the two fluorophores was extracted from it. The results of the experiment show the highest signal-to-noise ratio in the region just above the critical angle of 61.31º because of the higher signal intensity. However, for experiments where bulk exclusion is important, we observe the highest signal-to-bulk ratio at angles above 68˚ for a glass-water interface. Understanding the angular dependence on the sensitivity of a SAF biosensor enables tuning the collection angles towards specific applications and could lead to the creation of smaller, more sensitive devices.

The last decade has witnessed a rapid growth of nanoscale-oriented biosensors that becomes one of the most promising and rapidly growing areas of modern research. Despite significant advancements in conception of such biosensors, most are based at evaluation of molecular, or protein interactions. It is however becoming increasingly evident that functionality of a living system does not reside in genome or in individual proteins, as no real biological functionality is expressed at these levels. Instead, to comprehend the true functioning of a biological system, it is essential to understand the integrative physiological behavior of the complex molecular interactions in their natural environment and precise spatio-temporal topology. In this contribution we therefore present a new concept for creation of biosensors, bio-inspired from true functioning of living cells, while monitoring their endogenous fluorescence, or autofluorescence.

The programmed assembly of gold nanoparticles on DNA templates allows the design of nanostructures with optical properties that directly depend on the morphology of a biochemical scaffold. Indeed, the nanometer-scale sensitivity of plasmon coupling allows the translation of minute conformational changes into macroscopic optical signals. In particular, we showed that it is possible to design DNA-linked gold nanoparticle dimers that act as nanoscale actuators: nanostructures that change conformation under an external stimulus such as the hybridization of a specific DNA strand (L. Lermusiaux et al, ACS Nano 6, 10992 (2012)). Importantly, we are able to monitor, on a simple color camera, the conformational changes of a single DNA-assembled gold particle dimer as its gap is reduced from 20 nm to 1 nm when varying the local ionic strength (L. Lermusiaux et al, ACS Nano 9, 978 (2015)). We will also discuss recent results on the use of 3D DNA origamis as scaffolds for the assembly of plasmonic nanostructures, demonstrating that the conformation of the origami can be correlated to single nanostructure spectroscopy measurements. The flexibility of these biochemical templates opens exciting perspectives for the optical sensing of specific physicochemical stimuli that actively modify their 3D conformation, such as short biomolecules, specific cations or organic trace elements in water.

Surface Plasmon Resonance (SPR) is currently being widely studied as it exhibits sensitive optical properties to changes in in the refractive index of the surrounding medium. As novel devices using SPR have been developing rapidly there is a necessity to develop models and simulation environments that will allow for continued development and optimization of these devices. A biological sensing device of interest is the Plasmon FET which has been proven experimentally to have a limit of detection (LOD) of 20pg/ml while being immune to the absorption of the medium. The Plasmon FET is a metal-semiconductor-metal detector which employ functionalized gold nanostructures on a semi-conducting layer. This direct approach has the advantages of not requiring readout optics reducing size and allowing for point-of -care measurements. Using Lumerical FDTD and Device numerical solvers, we can report an advanced simulation environment illustrating several key sensor specifications including LOD, resolution, sensitivity, and dynamic range, for a variety of biological markers providing a comprehensive analysis of a Direct Plasmon-to-Electric conversion device designed to function with colored mediums (eg.whole blood). This model allows for the simulation and optimization of a plasmonic sensor that already o ers advantages in size, operability, and multiplexing-capability, with real time monitoring.

Using polarized light in medical imaging is a valuable tool for diagnostic purposes since light traveling through scattering tissues such as skin, blood, or cartilage may be subject to changes in polarization. We present a new detection scheme and sensor that allows for directly measuring the polarization of light electronically using a plasmonic sensor. The sensor we fabricated consists of a plasmonic nano-grating that is embedded in a Wheatstone circuit. Using resistive losses induced by optically excited plasmons has shown promise as a CMOScompatible plasmonic light detector. Since the plasmonic response is sensitive to polarization with respect to the grating orientation, measuring the resistance change under incident light supplies a direct electronic measure of the polarization of light without polarization optics. Increased electron scattering introduced by plasmons in an applied current results in a measurable decrease in electrical conductance of a grating, allowing a purely electronic readout of a plasmonic excitation. Accordingly, because of its plasmonic nature, such a detector is dependent on both the wavelength and polarization of incident light with a response time limited by the surface plasmon lifetime.

Photonics has become critical to life sciences. However, the field is far from benefiting fully from photonics' capabilities. Today, bulky and expensive optical systems dominate biomedical photonics, even though robust optical functionality can be realized cost-effectively on single photonic integrated circuits (PICs). Such chips are commercially available mostly for telecom applications, and at infrared wavelengths. Although proof-of-concept demonstrations for PICs in life sciences, using visible wavelengths are abundant, the gating factor for wider adoption is limited in resource capacity. Two European pilot lines, PIX4life and PIXAPP, were established to facilitate European R and D in biophotonics, by helping European companies and universities bridge the gap between research and industrial development. Through creation of an open-access model, PIX4life aims to lower barriers to entry for prototyping and validating biophotonics concepts for larger scale production. In addition, PIXAPP enables the assembly and packaging of photonic integrated circuits.

We describe recent research in which we explored biologically relevant logic gates using gold nanoparticles (GNPs) conjugated to fluorophores and tracing the results remotely by time-domain fluorescence lifetime imaging microscopy (FLIM).

GNPs have a well-known effect on nearby fluorophores in terms of their fluorescence intensity (FI – increase or decrease) as well as fluorescence lifetime (FLT). We have designed a few bio-switch systems in which the FLIMdetected fluorescence varies after biologically relevant stimulation. Some of our tools include fluorescein diacetate (FDA) which can be activated by either esterases or pH, peptide chains cleavable by caspase 3, and the polymer polyacrylic acid which varies in size based on surrounding pH. After conjugating GNPs to chosen fluorophores, we have successfully demonstrated the logic gates of NOT, AND, OR, NAND, NOR, and XOR by imaging different stages of activation. These logic gates have been demonstrated both in solutions as well as within cultured cells, thereby possibly opening the door for nanoparticulate in vivo smart detection.

While these initial probes are mainly tools for intelligent detection systems, they lay the foundation for logic gates functioning in conjunction so as to lead to a form of in vivo biological computing, where the system would be able to release proper treatment options in specific situations without external influence.

For many applications in life sciences, the biologically relevant information is probed by means of visible light. Many of the critical optical components have, unfortunately, still a large footprint and heavy price tag. Silicon nitride integrated waveguide optics –allowing for complex routing schemes of visible light across a chip– assumes a promi-nent role in the progressing miniaturization of optical devices. However, in order to have the light in the chip interro-gate a distant biological entity, diffraction gratings have to be used to couple light out of the chip. Ideally, all the light from a waveguide would be coupled out into a beam with a predefined polarization, phase, and intensity profile. As such they should be able to produce any functional beam that is typically prepared by free space optical components. For a standard, linear grating an exponential intensity decay is observed along the grating, i.e., more light is coupled out at the start than at the end. Here, we present a specially designed metasurface that is able to deliver highly uniform illumination escaping the photonics chip in a collimated beam at a predesigned angle. Because of its integrated nature, a component like this is highly relevant for the miniaturization of, e.g., flow cytometry applications. We therefore include microfluidic chan-nels on top of the photonics chip and demonstrate the cytometric capabilities with fluorescent polystyrene beads. The opto-fluidic chips are processed in a CMOS pilot line. Our work demonstrates the potential of integrated visible pho-tonics and flat optics for life science applications.

The detection sensitivity of surface plasmon resonance (SPR) biosensors has been improved by employing colocalization of spatial distribution of electromagnetic near-fields and detection molecules. We have used plasmon nanolithography to achieve light-matter colocalization on triangular nanoaperture arrays and optimized array configurations to improve colocalization efficiency. Streptavidin-biotin interactions were measured to validate the concept. It was confirmed that colocalized distributions of target binding and localized near-fields produced larger optical detection sensitivity. The colocalized detection was also shown to come with wider dynamic range than noncolocalized detection. The effective limit-of-detection of colocalized measurements was on the order of 30 pM. The colocalized detection sensitivity was estimated to be below 1 fg/mm² in a 100-nm deep evanescent area, an enhancement by more than three orders of magnitude over conventional SPR sensor.

Optical methods for extracting properties of tissues are commonly used. These methods are non-invasive, cause no harm to the patient and are characterized by high speed. The human tissue is a turbid media hence it poses a challenge for different optical methods. In addition the analysis of the emitted light requires calibration for achieving accuracy information. Most of the methods analyze the reflected light based on their phase and amplitude or the transmitted light. We suggest a new optical method for extracting optical properties of cylindrical tissues based on their full scattering profile (FSP), which means the angular distribution of the reemitted light. The FSP of cylindrical tissues is relevant for biomedical measurement of fingers, earlobes or pinched tissues. We found the iso-pathlength (IPL) point, a point on the surface of the cylinder medium where the light intensity remains constant and does not depend on the reduced scattering coefficient of the medium, but rather depends on the spatial structure and the cylindrical geometry. However, a similar behavior was also previously reported in reflection from a semi-infinite medium. Moreover, we presented a linear dependency between the radius of the tissue and the point's location. This point can be used as a self-calibration point and thus improve the accuracy of optical tissue measurements. This natural phenomenon has not been investigated before. We show this phenomenon theoretically, based on the diffusion theory, which is supported by our simulation results using Monte Carlo simulation.

Light reflectance and transmission from soft tissue has been utilized in noninvasive clinical measurement devices such as the photoplethysmograph (PPG) and reflectance pulse oximeter. Most methods of near infrared (NIR) spectroscopy focus on the volume reflectance from a semi-infinite sample, while very few measure transmission. However, since PPG and pulse oximetry are usually measured on tissue such as earlobe, fingertip, lip and pinched tissue, we propose examining the full scattering profile (FSP), which is the angular distribution of exiting photons. The FSP provides more comprehensive information when measuring from a cylindrical tissue. In our work we discovered a unique point, that we named the iso-pathlength (IPL) point, which is not dependent on changes in the reduced scattering coefficient (µs’). This IPL point was observed both in Monte Carlo (MC) simulation and in experimental tissue mimicking phantoms. The angle corresponding to this IPL point depends only on the tissue geometry. In the case of cylindrical tissues this point linearly depends on the tissue diameter. Since the target tissues for clinically physiological measuring are not a perfect cylinder, in this work we will examine how the change in the tissue cross section geometry influences the FSP and the IPL point. We used a MC simulation to compare a circular to an elliptic tissue cross section. The IPL point can serve as a self-calibration point for optical tissue measurements such as NIR spectroscopy, PPG and pulse oximetery.

Theranostics is an emerging field, defined as combination of therapeutic and diagnostic capabilities in the same material. Nanoparticles are considered as an efficient platform for theranostics, particularly in cancer treatment, as they offer substantial advantages over both common imaging contrast agents and chemotherapeutic drugs. However, the development of theranostic nanoplatforms raises an important question: Is the optimal particle for imaging also optimal for therapy? Are the specific parameters required for maximal drug delivery, similar to those required for imaging applications? Herein, we examined this issue by investigating the effect of nanoparticle size on tumor uptake and imaging. Anti-epidermal growth factor receptor (EGFR)-conjugated gold nanoparticles (GNPs) in different sizes (diameter range: 20–120 nm) were injected to tumor bearing mice and their uptake by tumors was measured, as well as their tumor visualization capabilities as tumor-targeted CT contrast agent. Interestingly, the results showed that different particles led to highest tumor uptake or highest contrast enhancement, meaning that the optimal particle size for drug delivery is not necessarily optimal for tumor imaging. These results have important implications on the design of theranostic nanoplatforms.

Despite the significant improvement in the treatment paradigm of head and neck cancer, owing to advanced radiation techniques in combination with chemotherapy, resistance of tumors remains a critical problem, leading to poor outcomes and negative prognosis. In addition, chemotherapeutic agents result in severe systemic toxicity due to nonselective damaging of normal cells. Recently, nanoparticle-based approaches have gained broad attention for improving both radiation therapy and chemotherapy. In this study, we present a dual effect nanoplatform, consists of gold nanoparticles coated with glucose and cisplatin (CG-GNPs), which simultaneously acts as a radiosensitizer and as a carrier which specifically deliver cisplatin to head and neck tumor. Our CG-GNPs showed significant penetration into tumor cells and similar cellular toxicity as cisplatin alone. Moreover, in combination with radiation treatment, CG-GNPs led to greater tumor reduction than that of free cisplatin with radiation. Furthermore, our CG-GNPs also demonstrated highly efficient imaging capabilities, as they act as ideal tumor-targeted CT contrast agent. Therefore, this single nano-formulation is a promising theranostic agent that has the potential to increase the antitumor effect and allow imaging guided therapy.

The luminescence spectra of upconversion nanoparticles (UCNPs) and ZnCdS nanoparticles (ZnCdSNPs) were measured and analyzed in a wide temperature range: from room to human body and further to a hyperthermic temperature resulting in tissue morphology change. The results show that the luminescence signal of UCNPs and ZnCdSNPs placed within the tissue is reasonably good sensitive to temperature change and accompanied by phase transitions of lipid structures of adipose tissue. The most likely that the multiple phase transitions are associated with the different components of fat cells, such as phospholipids of cell membrane and lipids of fat droplets. In the course of fat cell heating, lipids of fat droplet first transit from a crystalline form to a liquid crystal form and then to a liquid form, which is characterized by much less scattering. The results of phase transitions of lipids were observed as the changes in the slope of the temperature dependence of the intensity of luminescence of the film with nanoparticles embedded into tissue. The obtained results confirm a high sensitivity of the luminescent UCNPs and ZnCdSNPs to the temperature variations within thin tissue samples and show a strong potential for the controllable tissue thermolysis.

Cell-based therapies use living cells with therapeutic traits to treat various diseases. This is a beneficial alternative for diseases that existing medicine cannot cure efficiently. However, inconsistent results in clinical trials are preventing the advancement and implementation of cell-based therapy. In order to explain such results, there is a need to discover the fate of the transplanted cells. To answer this need, we developed a technique for noninvasive in vivo cell tracking, which uses gold nanoparticles as contrast agents for CT imaging. Herein, we investigate the design principles of this technique for intramuscular transplantation of therapeutic cells. Longitudinal studies were performed, demonstrating the ability to track cells over long periods of time. As few as 500 cells could be detected and a way to quantify the number of cells visualized by CT was demonstrated. This cell-tracking technology has the potential to become an essential tool in pre-clinical studies as well as in clinical trials and advance cell therapy.

Cell-to-cell communication system involves Exosomes, small, membrane-enveloped nanovesicles. Exosomes are evolving as effective therapeutic tools for different pathologies. These extracellular vesicles can bypass biological barriers such as the blood-brain barrier, and can function as powerful nanocarriers for drugs, proteins and gene therapeutics. However, to promote exosomes' therapy development, especially for brain pathologies, a better understanding of their mechanism of action, trafficking, pharmacokinetics and bio-distribution is needed. In this research, we established a new method for non-invasive in-vivo neuroimaging of mesenchymal stem cell (MSC)-derived exosomes, based on computed tomography (CT) imaging with glucose-coated gold nanoparticle (GNP) labeling. We demonstrated that the exosomes were efficiently and directly labeled with GNPs, via an energy-dependent mechanism. Additionally, we found the optimal parameters for exosome labeling and neuroimaging, wherein 5 nm GNPs enhanced labeling, and intranasal administration produced superior brain accumulation. We applied our technique in a mouse model of focal ischemia. Imaging and tracking of intranasally-administered GNP-labeled exosomes revealed specific accumulation and prolonged presence at the lesion area, up to 24 hrs. We propose that this novel exosome labeling and in-vivo neuroimaging technique can serve as a general platform for brain theranostics.

Characterizing different pathological states in the cellular level with a high throughput diagnostic tool is one of the main interests today. In previously works, we demonstrated how the frequency domain (FD) fluorescence lifetime imaging microscopy (FLIM) technique could be utilized to implement that in variety of examples. Among them was to classify between different chromosomal abnormalities in patients with b-cell chronic lymphocytic leukemia (B-CLL) and between metastatic cells and inflammation cells in the cerebral spinal fluid of patients with Medulloblastoma. This research describes the use of FD-FLIM system to differentiate between patients diagnosed without any disease (controls) that showed a normal median FLT (2.65±0.11ns) and patients diagnosed with inflammation (viruses and bacteria) that showed a prolong median FLT and a larger distribution (3.18±0.44ns in viruses and 3.28±0.45ns). The study group of this research included 43 samples divided into 4 groups: 9 samples diagnosed with different types of bacteria, 16 samples diagnosed with different types of viruses, 12 samples diagnosed with no any bacteria or virus and 5 samples diagnosed without any disease that served as controls.

Furthermore, we studied a group of patients without detection of inflammation that were sick. We found that this group was divided into two groups; one group had the same median FLT as the controls, and the other group had the same median FLT as the inflammatory patients. As a result, we believe the FD-FLIM system can suggest a faster and more accurate diagnostic technique than the methods used today. The correlations of the FLT distribution pattern with the different groups are presented.

Significant effort has been invested into understanding the dynamics of protein adsorption on surfaces, in particular to predict protein behavior at the specialized surfaces of biomedical technologies like hydrogels, nanoparticles, and biosensors. Recently, the application of fluorescent single molecule imaging to this field has permitted the tracking of individual proteins and their stochastic contribution to the aggregate dynamics of adsorption. However, the interpretation of these results is complicated by (1) the finite time available to observe effectively infinite adsorption timescales and (2) the contribution of photobleaching kinetics to adsorption kinetics. Here, we perform a protein adsorption simulation to introduce specific survival analysis methods that overcome the first complication. Additionally, we collect single molecule residence time data from the adsorption of fibrinogen to glass and use survival analysis to distinguish photobleaching kinetics from protein adsorption kinetics.

Extracting optical parameters of turbid medium (e.g. tissue) by light reflectance signals is of great interest and has many applications in the medical world, life science, material analysis and biomedical optics. The reemitted light from an irradiated tissue is affected by the light's interaction with the tissue components and contains the information about the tissue structure and physiological state. In this research we present a novel noninvasive nanophotonics technique, i.e., iterative multi-plane optical property extraction (IMOPE) based on reflectance measurements. The reflectance based IMOPE was applied for tissue viability examination, detection of gold nanorods (GNRs) within the blood circulation as well as blood flow detection using the GNRs presence within the blood vessels. The basics of the IMOPE combine a simple experimental setup for recording light intensity images with an iterative Gerchberg–Saxton (G-S) algorithm for reconstructing the reflected light phase and computing its standard deviation (STD). Changes in tissue composition affect its optical properties which results in changes in the light phase that can be measured by its STD. This work presents reflectance based IMOPE tissue viability examination, producing a decrease in the computed STD for older tissues, as well as investigating their organic material absorption capability. Finally, differentiation of the femoral vein from adjacent tissues using GNRs and the detection of their presence within blood circulation and tissues are also presented with high sensitivity (better than computed tomography) to low quantities of GNRs (<3 mg).

Giant unilamellar vesicles (GUVs) are well-established model systems for studying lipid packing and membrane dynamics. With sizes larger than 1 μm, GUVs are easily observable using optical microscopy. Gold nanoparticles (AuNPs) are well known for their biocompatibility and such biomedical applications in drug and gene delivery as well as medical diagnostics and therapeutics. On the other hand, silver nanoparticles (AgNPs) have long been known for their potent antimicrobial and anti-inflammatory effects for such applications as wound dressing and biomedical implants. In this work, we employed the dark-field microscopy (CytoViva Inc.) to study the interactions between AuNPs/AgNPs and GUVs, respectively. The GUVs used in this study were prepared with 1,2 dimyristoyl-sn-glycero-3-phosphocholine (DMPC) as well as cholesterol (chol) at various mol% concentrations (0, 10, 20, 30, 40%). The electroformed GUVs were allowed to incubate with gold or silver nanoparticles of various sizes (between 10 and 100 nm) for 2 hrs before microscopic examination. The experiment has shown that the size of nanoparticles is a critical factor that determines the penetration rate. In addition, the membrane rigidity increases with the molar concentration of cholesterol hence making the NP penetration more difficult. Comparative studies have been made between AuNPs and AgNPs in regard to NP penetration and loading rate as well as the morphological changes induced in GUVs. This work aims to better understand the mechanisms of AuNP/AgNP and membrane interactions for their respective future applications in nanomedicine and nanotechnology.

Pancreatic islet transplantation is a promising approach of providing insulin in type 1 diabetes. One strategy to protect islets from the host immune system is encapsulation within a porous biocompatible alginate membrane. This encapsulation provides mechanical support to the cells and allows selective diffusion of oxygen, nutrients and insulin while blocking immunoglobulins. These hydrogels form by diffusion of calcium ions into the polymer network and therefore they are highly sensitive to environmental changes and fluctuations in temperature. We investigated the effects of gel concentration, crosslinking time and ambient conditions on material permeability, volume, and rigidity, all of which may change the immunoisolating characteristics of alginate. To measure diffusion coefficient as a method to capture structural changes we studied the diffusion of fluorescently tagged dextrans of different molecular weight into the midplane of alginate microcapsules, the diffusion coefficient is then calculated by fitting observed fluorescence dynamics to the mathematical solution of 1-D diffusion into a sphere. These measurements were performed after incubation in different conditions as well as after an in vivo experiment in six immunocompetent mice for seven days. Additionally, the changes in gel volume after incubation at different temperatures and environmental conditions as well as changes in compression modulus of alginate gels during crosslinking were investigated. Our result show that increase of polymer concentration and crosslinking time leads to a decrease in volume and increase in compression modulus. Furthermore, we found that samples crosslinked and placed in physiological environment, experience an increase in volume. As expected, these volume changes affect diffusion rates of fluorescent dextrans, where volume expansion is correlated with higher calculated diffusion coefficient. This observation is critical to islet protection since higher permeability due to the expansion in vivo may lead to increased permeability to immunoglobulins. Capsules from the in vivo study showed similar volume expansion and increased permeability, indicating our in vitro assay is a good predictor of volume change in vivo.

Background subtraction of knee MRI images has been performed, followed by edge detection through canny edge detector. In order to avoid the discontinuities among edges, Daubechies-4 (Db-4) discrete wavelet transform (DWT) methodology is applied for the smoothening of edges identified through canny edge detector. The approximation coefficients of Db-4, having highest energy is selected to get rid of discontinuities in edges. Hough transform is then applied to find imperfect knee locations, as a function of distance (r) and angle (θ). The final outcome of the linear Hough transform is a two-dimensional array i.e., the accumulator space (r, θ) where one dimension of this matrix is the quantized angle θ and the other dimension is the quantized distance r. A novel algorithm has been suggested such that any deviation from the healthy knee bone structure for diseases like osteoarthritis can clearly be depicted on the accumulator space.