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Summary: Emerging evidence suggests that crosstalk between glioma cells and the brain microenvironment may influence brain tumor growth. To date, known reciprocal interactions among these cells have been limited to the release of paracrine factors. Combining a genetic strategy with longitudinal live imaging, we find that individual gliomas communicate with distinct sets of non-glioma cells, including glial cells, neurons, and vascular cells. Transfer of genetic material is achieved mainly through extracellular vesicles (EVs), although cell fusion also plays a minor role. We further demonstrate that EV-mediated communication leads to the increase of synaptic activity in neurons. Blocking EV release causes a reduction of glioma growth in vivo. Our findings indicate that EV-mediated interaction between glioma cells and non-glioma brain cells alters the tumor microenvironment and contributes to glioma development. : Non-glioma cells form a unique tumor microenvironment and are critical for glioma progression. Gao et al. find that individual gliomas communicate with distinct sets of non-glioma cells, including glial cells, neurons, and vascular cells. Transfer of genetic material is achieved mainly through extracellular vesicles (EVs). Keywords: glioma, glia, astrocyte, interaction, extracellular vesicle, tumor microenvironment, cell fusion, neuron, exosome

We report fabrication and implementation of fiber-tip probes with localized surface plasmon resonators for in situ biosensing applications. Gold nano-void arrays, which act as localized surface plasmon resonators were fabricated on ordinary glass coverslips and mounted on cleaved tips of multi-mode optical fibers. Plasmon resonance peak wavelength and depth are functions of gold nano-void array parameters. Array design parameters were optimized using numerical simulations to attain maximum resonance depth and peak in the near infra-red region of the optical spectrum. Sensitivity of the fiber-tip localized surface plasmon resonators to quantify change in refractive index is measured to be 535.2 nm/RIU. Proof-of-principle experimental results of biomolecular interaction analysis with functionalized fiber-tip resonators are presented.

Electrophysiology techniques are the gold standard in neuroscience for studying functionality of a single neuron to a complex neuronal network. However, electrophysiology techniques are not flawless, they are invasive nature, procedures are cumbersome to implement with limited capability of being used as a high-throughput recording system. Also, long term studies of neuronal functionality with aid of electrophysiology is not feasible. Non-invasive stimulation and detection of neuronal electrical activity has been a long standing goal in neuroscience. Introduction of optogenetics has ushered in the era of non-invasive optical stimulation of neurons, which is revolutionizing neuroscience research. Optical detection of neuronal activity that is comparable to electro-physiology is still elusive. A number of optical techniques have been reported recording of neuronal electrical activity but none is capable of reliably measuring action potential spikes that is comparable to electro-physiology. Optical detection of action potential with voltage sensitive fluorescent reporters are potential alternatives to electrophysiology techniques. The heavily rely on secondary reporters, which are often toxic in nature with background fluorescence, with slow response and low SNR making them far from ideal. The detection of one shot (without averaging)-single action potential in a true label-free way has been elusive so far. In this report, we demonstrate the optical detection of single neuronal spike in a cultured mammalian neuronal network without using any exogenous labels. To the best of our knowledge, this is the first demonstration of label free optical detection of single action potentials in a mammalian neuronal network, which was achieved using a high-speed phase sensitive interferometer. We have carried out stimulation and inhibition of neuronal firing using Glutamate and Tetrodotoxin respectively to demonstrate the different outcome (stimulation and inhibition) revealed in optical signal. We hypothesize that the interrogating optical beam is modulated during neuronal firing by electro-motility driven membrane fluctuation in conjunction with electrical wave propagation in cellular system.

Quantitative analysis of Raman spectra using surface-enhanced Raman scattering (SERS) nanoparticles has shown the potential and promising trend of development in in vivo molecular imaging. Partial least square regression (PLSR) methods have been reported as state-of-the-art methods. However, the approaches fully rely on the intensities of Raman spectra and can not avoid the influences of the unstable background. In this paper we design a new continuous wavelet transform based PLSR (CWT-PLSR) algorithm that uses mixing concentrations and the average CWT coefficients of Raman spectra to carry out PLSR. We elaborate and prove how the average CWT coefficients with a Mexican hat mother wavelet are robust representations of Raman peaks, and the method can reduce the influences of unstable baseline and random noises during the prediction process. The algorithm was tested using three Raman spectra data sets with three cross-validation methods in comparison with current leading methods, and the results show its robustness and effectiveness.

Brain tumor cells remain highly resistant to radiation and chemotherapy, particularly malignant and secondary cancers. In this study, we utilized microchannel devices to examine the effect of a confined environment on the viability and drug resistance of the following brain cancer cell lines: primary cancers (glioblastoma multiforme and neuroblastoma), human brain cancer cell lines (D54 and D54-EGFRvIII) and genetically modified mouse astrocytes (wild type, p53−/−, p53−/− PTEN−/−, p53−/− Braf and p53−/− PTEN−/− Braf). We found that loss of PTEN combined with Braf activation resulted in higher viability in narrow microchannels. In addition, Braf conferred increased resistance to the microtubule-stabilizing drug Taxol in narrow confinement. Similarly, survival of D54-EGFRvIII cells was unaffected following treatment with Taxol, whereas the viability of D54 cells was reduced by 75% under these conditions. Taken together, our data suggests key targets for anticancer drugs based on cellular genotypes and their specific survival phenotypes during confined migration.

Detecting cellular activity in sub-millisecond timescale and micrometer resolution without using invasive means has been a long standing goal in the study of cellular networks. Here, we have employed phase sensitive low coherence interferometry for detecting optogenetically stimulated activity of cells. Nanoscale changes in optical path length (due to change in refractive index and changes in cell thickness) occur when cells are activated, which we aim to detect by phase sensitive low coherence interferometry. A low coherence interferometry and patch-clamp electrophysiology systems were integrated with an inverted fluorescence microscope. Blue laser beam was coupled to the electrophysiology-interferometric detection system for optogenetic stimulation. The phase-sensitive measurements were carried out on Channelrhodopsin-2 sensitized cells (identified by YFP fluorescence) as well as control cells in reflection mode for different intensities and exposures of optogenetic stimulation beam. This method offers good temporal and spatial resolution without using exogenous labeling. Results of studies on all optical stimulation and detection of cellular activity will be presented. Interpretation of the optical activity signals will be discussed in context with changes in cell physiology during stimulation. We will also discuss the potential sources of various artifacts in optical/electrical detection of cellular activity during optical stimulation.

Here Phase Sensitive-Spectral Domain Optical Coherence Tomography is used to detect cellular activity in HEK and primary neurons in an all optical, label free manner in sub-millisecond timescale and micrometer resolution without using invasive means.

Plasmonic properties of gold nanovoid array substrates for fiber-based surface-enhanced Raman scattering (SERS) sensing are studied numerically and experimentally. In the nanovoid arrays, each void has openings on both sides, bottom hole facing the fiber tip for introducing incident light and collecting scattered light and the top hole exposed to the analyte solution for interrogating analyte molecules in the voids. Electromagnetic field modes are confined strongly in and around these nanovoids, acting as localized plasmon resonators. The enhanced electric field extends hundreds of nanometers into the voids, resulting in a large SERS-active zone several orders of magnitude larger than nanoparticle-based structures. The effect of structural parameters of the nanovoid arrays, including void diameter, Au film thickness, and bottom hole diameter, on electric field confinement in the voids is investigated using three-dimensional finite difference time domain simulation. Au nanovoid arrays are fabricated using a scalable, inexpensive nanosphere lithography fabrication method. The largest SERS signal is realized by tuning the localized plasmon resonance peak of Au nanovoid arrays to the laser excitation wavelength. Multiplexed detection capability with the fiber-based SERS sensor using Au nanovoid arrays is demonstrated by measuring the Raman spectrum of a mixture solution of diethylthiatricarbocyanine and crystal violet.

Surface enhanced Raman spectroscopy (SERS) is a promising molecular imaging modality capable of simultaneously detecting multiple molecular biomarkers. With the biocompatibility and functionalizability of Au, Au-nanoparticle based Raman tags possess the potential for in vivo SERS cancer biomarker detection. Here, we report the large scale synthesis of a new type of Au nanoparticles, Porous Hollow Au Nanoparticles (PHAuNPs), and demonstrate their potential application as SERS imaging tags. PHAuNPs feature a sub-20 nm porous shell and a 50 nm void core. Such unique morphology enables them to strongly absorb and scatter near infrared lights due to the surface plasmon resonant effect of Au. This makes them particularly suitable for in vivo applications, where NIR wavelengths are considered as a ‘clear window’ for deeper penetration of light. The construction and characterization of PHAuNP-based Raman nanotag, including attachment of Raman dye, pegylation and their stability, are described. Cytotoxicity of Raman nanotags are tested using the radioactive [3H]thymidine incorporation method. The results show that pegylated Raman nanotags are stable and non-toxic and can potentially be used for in vivo applications.

With the latest development of Surface-Enhanced Raman Scattering (SERS) technique, quantitative analysis of Raman spectra has shown the potential and promising trend of development in vivo molecular imaging. Partial Least Squares Regression (PLSR) is state-of-the-art method. But it only relies on training samples, which makes it difficult to incorporate complex domain knowledge. Based on probabilistic Principal Component Analysis (PCA) and probabilistic curve fitting idea, we propose a probabilistic PLSR (PPLSR) model and an Estimation Maximisation (EM) algorithm for estimating parameters. This model explains PLSR from a probabilistic viewpoint, describes its essential meaning and provides a foundation to develop future Bayesian nonparametrics models. Two real Raman spectra datasets were used to evaluate this model, and experimental results show its effectiveness.

Biomedical Engineering Department, University of Texas of Austin, TexasBackground and Objectives: We describe a methodol-ogytorecordspatialvariation ofrefractiveindexofporcinerenal artery using differential phase optical coherencemicroscopy (DP-OCM).Study Design/Materials and Methods: The DP-OCMprovidesquantitativemeasurementofthinspecimenphaseretardationandrefractiveindexbymeasuringopticalpath-lengthchangesontheorderofafewnanometersandwithalateralresolutionof3mm.TheDP-OCMinstrumentationisan all-fiber, dual-channel Michelson interferometer con-structed using a polarization maintaining (PM) fiber.Results: Two-dimensional en face dual-channel phaseimages are taken over a 150 200 mm region on amicroscopic slide, and the images are reconstructed byplotting a two-dimensional refractive index map as theOCM beam is moved across the sample.Conclusions: Because the DP-OCM can record transientchangesintheopticalpath-length,thesystemmaybeusedto record quantitative optical path-length alterations oftissue in response to various stimuli. A fiber-based DP-OCM may have the potential to substantially improve invivo imaging of individual cells for a variety of clinicaldiagnostics, and monitoring applications. Lasers Surg.Med. 38:955–959, 2006. 2006 Wiley-Liss, Inc.Key words: phase sensitive optical coherence micro-scopy; refractive index; relative phase retardationINTRODUCTIONOptical coherence tomography (OCT) has emerged as animportantopticalimagingmodalityinnoninvasivemedicaldiagnostics. OCT generates high-resolution images byutilizing the cross correlation between light backscatteredfrom the sample and the reference reflector. Varioustechnical approaches have been developed to improveOCT spatial resolution, imaging acquisition rate [1,2],andimagequality[3,4].Mostofthesetechniquesusefringeamplitude or a combination of fringe amplitude andpolarization information in reflected light [5]. Magnitudeof back-reflected light is dependent on the local refractiveindex gradients of the structural components comprisingthe test specimen. Polarization information basicallyreveals different phase retardation, caused by a tissue,between two orthogonal polarization states of light.Knowledge of local refractive index variations in biologi-cal tissue is not only important in tissue optics but also amajor factor that determines fringe signal variation inconventionalOCT.Averagerefractiveindexoverrelativelyhomogeneous tissue medium has been measured byTearney et al.[6] and Knu¨ttel et al.[7]. The basic approachpresented by Tearney et al. [6] was to adjust the referencemirror for maximum signal at a particular focus positionwithinatissuewhereasKnu¨tteletal.[7]adaptedamovingfiber tip/collimating lens in the sample path in order totrace maximum signal variation. Two-dimensional bire-fringence mapping of porcine myocardium tissue wasreported [6]. The DP-OCM is capable of measuring opticalpath-length changes on the order of a few nanometers ( 5nm) [8] so that it can be applied to image two-dimensionallocal refractive index variation of thin specimens.This article presents a method to image refractive indexdistribution over a tissue specimen processed by standardhistology sectioning methods and fixated on a microscopeslide.MATERIALS AND METHODSDP-OCM Instrumentation and Signal ProcessingDetailsoftheDP-OCMusedforrefractiveindexprofilingcan be found in references [8,9]. A simplified schematicdiagram is shown in Figure 1. The DP-OCM system isessentially a dual-channel Michelson interferometer withtwo independent channels corresponding to orthogonalpolarization modes of a polarization maintaining (PM)optical fiber. At the interferometer input, light emittedfrom a broadband source (l

We present a novel fiber-optic spectral polarimetry instrument based on optical frequency domain interferometry to provide measurement of the Stokes spectra of collected light. Stokes spectra are encoded into discrete channels in the time delay domain. The instrument consists of a broadband swept laser source, calibrated wavemeter, two segments of polarization maintaining (PM) fiber, an analyzer and photo-receiver. Performance of the instrument is demonstrated by reconstruction of Stokes spectra for a variety of prepared incident polarization states.

We describe a differential phase low-coherence interferometric probe for non-invasive, quantitative imaging of photothermal phenomena in biological materials. Our detection method utilizes principles of optical coherence tomography with differential phase measurement of interference fringe signals. A dual-channel optical low-coherence probe is used to analyse laser-induced thermoelastic and thermorefractive effects in tissue with micrometre axial resolution and nanometre sensitivity. We demonstrate an application of the technique using tissue phantoms and ex-vivo tissue specimens of rodent dorsal skin.

Background and Objectives Tissue response to thermal, electrical, or chemical stimuli are important in the health and survival of tissue. We report experimental results to assess tissue response to various stimuli using a low coherence differential phase interferometer. Study Design/Materials and Methods The optical system utilized to measure tissue response is a novel fiber-based phase sensitive optical low coherence reflectometer (PS-OLCR). Inasmuch as the PS-OLCR works with back-reflected light, noninvasive sensing of tissue response to stimuli is possible. In addition to high lateral (∼10 μm) and longitudinal (∼10 μm) resolution, PS-OLCR can measure sub-wavelength changes in optical path-length (Angstrom/nanometer range) by extracting the phase difference between interference fringes in two channels corresponding to orthogonal polarization modes. Results When light spatially splits into two polarization states, precise analysis of surface topography or tissue surface response such as swelling or collapse are possible. Time resolved measurements of nanometer-scale path length changes in response to electrical and thermal stimuli are demonstrated using longitudinally delayed polarization channels. Conclusions Since PS-OLCR is a useful tool to detect ultra-small path length changes, the system has potential to aid scientists in investigating important phenomena in biomaterials and developing useful diagnostic and therapeutic imaging modalities. Applications include tissue surface profilometry, measurement of tissue, and cell response to various stimuli, high-resolution intensity and phase imaging. Lasers Surg. Med. 33:219–225, 2003. © 2003 Wiley-Liss, Inc.

We describe an optical technique for label-free detection of the action potential in cultured mammalian neurons. Induced morphological changes due to action potential propagation in neurons are optically interrogated with a phase sensitive interferometric technique. Optical recordings composed of signal pulses mirror the electrical spike train activity of individual neurons in a network. The optical pulses are transient nanoscale oscillatory changes in the optical path length of varying peak magnitude and temporal width. Exogenous application of glutamate to cortical neuronal cultures produced coincident increase in the electrical and optical activity; both were blocked by application of a Na-channel blocker, Tetrodotoxin. The observed transient change in optical path length in a single optical pulse is primarily due to physical fluctuations of the neuronal cell membrane mediated by a yet unknown electromechanical transduction phenomenon. Our analysis suggests a traveling surface wave in the neuronal cell membrane is responsible for the measured optical signal pulses.

The ability to diffusely infiltrate as single cells through the highly confined extracellular spaces of the brain parenchyma and along blood vessels is a pathological hallmark of Glioblastoma Multiforme (GBM), which makes these tumors surgically incurable. The fact that GBM cells are capable of moving through diverse brain microenvironments raises the possibility that GBM cells may possess multiple mechanisms for invasion. At present, little is known about how GBM cells gain traction and generate sufficient contractile forces to overcome the mechanical challenge of migrating through the confined interstitial spaces of the brain. The purpose of this study is to gain cellular, biochemical and biophysical insights into GBM migration. To accomplish this we generated 1-D micro-patterned arrays that approximate the topography of brain microvasculature and microfluidic devices that approximate brain interstitial spaces. High-resolution 3D time lapse imaging is then used to track actin, myosin-II and alpha-tubulin dynamics during cell migration. We found that the geometry of micro-patterned laminin lanes in the absence of chemotactic, hapatotactic or durotactic gradients, is sufficient to trigger sustained (integrin) adhesion-dependent migration that is driven by actin polymerization at the leading edge and supported by acto-myosin-II cable-like structures that extend along the ventral and lateral edges of spindle shaped cells. A similar pattern of adhesion-dependent (mesenchymal) migration was seen in 3D channels with aspect ratio (15x15um) larger than the cell. In confined (5x5um) channels, GBM cells adopted interchangeable modes of migration, waxing between blebbing and mesenchyal migration (approximately 50:50). Blebbing migration was characterized by acto-myosin-II cable-like structures that extended the length of the cell along both ventral and dorsal surfaces. In contrast, mesenchymal phase migration was characterized by a cytoskeleton organization pattern that resembled 1-D migration. Exposure of cells in confined channels to actin inhibitors (latrunculin B and CK-666) caused the cells to adopt a blebbing predominant mode of migration, while myosin-II inhibitor (blebbistatin) induced a mesenchymal predominant pattern. While neither class of inhibitors blocked migration as a single agent, the combination produced cell arrest. GBM cells were also able to migrate through confined channels in the absence of any ECM coating by adopting a blebbing exclusive pattern of migration. This observation raises the possibility that in the absence of integrin signaling, GBM cells can use hydrostatic pressure to gain traction in confined spaces. Taken together these results suggest that GBM can interchangeably adopt adhesion dependent and independent modes of migration as they encounter different sized ECM topography and variable 3-D confinement. Citation Format: James O. Nyagilo, Sara Picirrillo, Yuxiao Sun, Loan T. Bui, Nanda Regmi, Jamie L. Wright, Donald Thevalingam, Armin Soltan Zadi, Young-tae Kim, Charles Choung, Digant P. Dave, Robert Bachoo. Extracellular matrix geometry and 3D spatial confinement trigger diverse mechanisms of primary human glioblastoma cell migration. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr A54.

We describe a fast phase-sensitive frequency-multiplexed optical low-coherence reflectometer which is capable on measuring optical path-length changes on the order of a few nanometers (∼5 nm). Results are presented that demonstrate phase sensitivity and frequency multiplexing capability of the interferometer.

Surface-enhanced Raman spectroscopy (SERS) has been a routine method for the quantitative analysis of Nano-tags or biomarkers. The multivariate calibration (MC) model is normally used to reduce the bias from the inherent instability of Raman signals. To solve the more variables than observations, ill-conditioned problem within the MC model, latent variable regression (LVR) methods are usually used. In order to decide the optimized number of latent variables (LVs) used in the model, cross-validation methods are commonly used to test every possible number, and the one gives the minimum estimated error is returned as the optimized number. In this paper we present a new multi-component LVR together with a cross-validation framework to accelerate the time-consuming processes of optimizing number of LVs. It reduces the growth rate of the algorithms from O(k^2) to O(k), where k is the possible numbers of LVs. Experimental results show the estimated results of the two frameworks are equivalent and the running time of our new framework is evidently reduced.

Gold Surface-enhanced Raman Scattering (Au SERS) nano-particles in combination with Raman spectroscopy have occurred as a newly sensitive, non-invasive molecular imaging technology. The multiplexing capability enables the technology to detect and separate multiple biomarkers with picomolar sensitivity. In this study, we demonstrate the ability of Raman spectroscopy to separate different fingerprints of Au SERS nanotags. Quantitative analysis of Raman spectrum data usually faces the challenge as high dimensional variables with a low sample number. The commonly applied partial least squares (PLS) regression algorithms, including PLS2 and SIMPLS, can not avoid overfitting to small data sets. In this paper, we present a unified probabilistic PLSR model, called PPLSR, stemmed from the concepts of probabilistic principal component analysis (PPCA) and probabilistic canonical correlation analysis (PCCA) to identify the spectral fingerprints from the measured mixing Raman signals. This model partitions the observed variables into the systematic part governed by a few latent variables and the unrelated noise part controlling the uncertainty of data sets. As a general methodology, this provides a solid foundation to develop Bayesian nonparametrics models and helps to build more robust models. Experimental results of Raman spectrum data using up to five different types of Au SERS nanotags with different combinations and mixing ratios are shown. Quantitative analysis using the proposed model and comparison methods are given with two cross-validation methods.

The quantitative analysis of surface-enhanced Raman spectra using scattering nanoparticles has shown the potential and promising applications in in vivo molecular imaging. The diverse approaches have been used for quantitative analysis of Raman spectra information, which can be categorized as direct classical least squares models, full spectrum multivariate calibration models, selected multivariate calibration models, and latent variable regression (LVR) models. However, the working principle of these methods in the Raman spectra application remains poorly understood and a clear picture of the overall performance of each model is missing. Based on the characteristics of the Raman spectra, in this paper, we first provide the theoretical foundation of the aforementioned commonly used models and show why the LVR models are more suitable for quantitative analysis of the Raman spectra. Then, we demonstrate the fundamental connections and differences between different LVR methods, such as principal component regression, reduced-rank regression, partial least square regression (PLSR), canonical correlation regression, and robust canonical analysis, by comparing their objective functions and constraints. We further prove that PLSR is literally a blend of multivariate calibration and feature extraction model that relates concentrations of nanotags to spectrum intensity. These features (a.k.a. latent variables) satisfy two purposes: the best representation of the predictor matrix and correlation with the response matrix. These illustrations give a new understanding of the traditional PLSR and explain why PLSR exceeds other methods in quantitative analysis of the Raman spectra problem. In the end, all the methods are tested on the Raman spectra datasets with different evaluation criteria to evaluate their performance.

This study proposes a novel concept for a three-axis positioning and scanning platform that overcomes the existing gap in technology towards meeting the requirements for displacements, resolution, weight carrying capacity and velocity at smaller dimensions. The novelty of this work stems from the fact that our three-axis stage design utilizes only two actuators. This system was developed to meet the specific requirements needed for implementation of Multifunctional Image Guided Surgical (MIGS) platform. Mathematical model accounting for the open and closed loop operation of the stage was developed. The stage can provide displacements between 10-20mm in each axis, resolution of less than 10μm and scanning velocity in the range of 10-40mm/s. It can carry weights up to 10grams while meeting the desired requirements. Additionally, the stage has small footprint (50mm × 50mm × 34mm), modular design and extremely cost-effective fabrication. Integration of computer controlled three-axis stage with MIGS platform will provide the opportunity for conducting intricate surgical procedures using remote control or joystick. We demonstrate novel applications that became possible due to the development of this stage.

Quantitative analysis of Raman spectrum using Surface Enhanced Raman scattering (SERS) nanoparticles has shown the potential and promising trend of development in vivo molecular imaging. Because of the high dimension of Raman spectra and limited number of samples, latent variable regression methods, e.g. principal component regression (PCR), reduced-rank regression (RRR) and partial least squares (PLS), are commonly used. According to different criteria, these methods tend to seek different latent variables of the spectra data. For PCR and RRR, the latent variables tend to best represent the Raman spectra and best predict the concentrations. PLS balances the two criteria with an equal weight. We design a new continuum regression (NCR) method that uses a weight parameter ? to control the portion of each criterion in the objective function, and embraces RRR (? = 0), PLS2 (? = 1) and PCR (? = ?) as its special cases. The experimental results show that its performance is better than the other two continuum regression methods.

Quantitative analysis of Raman spectra using Surface Enhanced Raman scattering (SERS) nanoparticles has shown the potential and promising trend of development in vivo molecular imaging. Partial Least Square Regression (PLSR) methods are the state-of-the-art methods. But they rely on the whole intensities of Raman spectra and can not avoid the instable background. In this paper we design a new CWT-PLSR algorithm that uses mixing concentrations and the average continuous wavelet transform (CWT) coefficients of Raman spectra to do PLSR. The average CWT coefficients with a Mexican hat mother wavelet are robust representations of the Raman peaks, and the method can reduce the influences of the instable baseline and random noises during the prediction process. In the end, the algorithm is tested on three Raman spectrum data sets with three cross-validation methods, and the results show its robustness and effectiveness.

We have developed a high precision three axes scanning and positioning system for integration with Multifunctional Image Guided Surgical (MIGS) Platform. The stage integrates three main components: an optical coherence tomography (OCT) probe, laser scalpel and suction cup. The requirements for this stage were to provide scanning area of 400mm2, resolution of less than 10 microns and scanning velocity in the range of 10 - 40 mm/s. The stage was modeled using computer aided design software NX Unigraphics. In addition to the parameters mentioned above, additional boundary conditions for the stage were set as low volume and modularity. Optimized stage model was fabricated by using rapid prototyping technique that integrates low cost stepper motors, threaded rod drive train and a stepper motor controller. The EZ4axis stepper motor controller was able to provide 1/8th microstep resolution control over the motors, which met the criterion desired for the MIGS platform. Integration of computer controlled three-axis stage with MIGS platform provides the opportunity for conducting intricate surgical procedures using remote control or joystick. The device is image guided using the OCT probe and it is able to pin point any location requiring a laser scalpel incision. Due to the scanning capabilities, a high quality threedimensional image of the tissue topography is obtained which allows the surgeon to make a confident decision of where to apply the laser scalpel and make an incision.

A simple closed form expression for a sum rule is derived for quantum mechanical systems where the effective mass varies across the structure. It is a generalization of the Thomas-Kuhn sum rule for an effective mass which is independent of position.

Raman spectroscopy has been one of the most sensitive techniques widely used in chemical and pharmaceutical material identification research ever since it is invented based on Raman scattering theory, because of the fingerprints property of Raman signals to different materials. With the latest development of surface enhanced Raman scattering (SERS) nanoparticles, Raman spectroscopy is now used in more and more quantitative analysis applications. But due to the unavoidable instable problem of Raman spectroscopy signal, as well as the high signal dimension and small sample number problem, it is badly in need of a robust and accurate signal quantitative analysis method. Based on Partial Least Square Regression (PLSR) method, Probabilistic PCA and Probabilistic curve-fitting idea, we propose a new Probabilistic-PLSR (PPLSR) model. It explains PLSR from a probabilistic viewpoint and deeply describes the physical meaning of PLSR model. It is a solid foundation to develop more robust and accurate probabilistic PLSR models with Bayesian model in order to solve the over-fitting problem. And since this model adds a regularization term in the matrix of regression coefficients, the estimated result is more robust than PLSR model. We also provide an EM Algorithm to estimate the parameters of the model from sample data. To take fully use of the valuable data, we design two experiments, leave-one out and cross-validation-on-average-signal, on one real Raman spectroscopy signal data set. By comparing with results from traditional Least Square (LS) method and traditional PLSR, we demonstrate PPLSR is more robust and accurate.

We theoretically calculate the electric field induced refractive change index for the ground to first excited state transition in the conduction band of asymmetrical quantum wells. We show that there is a significant change in stark shifts and the refractive index with asymmetry potential. Comparing the results for a square, graded, and step well, it is found that larger refractive index change can be achieved in a step well by a proper choice of the step potential.

Raman spectroscopy has been one of the most sensitive techniques widely used in chemical and pharmaceutical research. With the latest development of surface enhanced Raman scattering (SERS) nanoparticles, the application now can be extended to bioimaging and biosensing. In this study, we demonstrate the ability of Raman spectroscopy to separate multiple spectral fingerprints using Raman nanotags after injection. The competence will further be used as functional agents for diagnostic molecular imaging applications. In this paper, a machine learning method is proposed to estimate the mixing ratios of each source signal from a mixture signal. The method first decomposes the training mixture signal matrix into a number of components and meanwhile keeps the maximum linear relationship between the new coordinate and ground truth ratio matrix. Then a regression coefficient matrix is formed by the component matrix. Traditional regression methods provide poor decomposition results due to various factors in sample preparation and machine operation that lead to the stochastic nature of Raman spectrum. The robustness of the proposed method was compared with least square and weighted least square methods.

A highly sensitive technique for detection and quantification of protein-analyte binding interaction using spectral domain low coherence interferometric technique is implemented. Using this technique, it is possible to study real time interactions by quantifying the changes in optical path length caused by protein-analyte binding. The technique does not require elaborate sensor preparation and calibration. Any coverslip with certain thickness and capability to bind protein can work using this technique. For proof of principle, the interaction between IgG antigen and anti-IgG was studied and quantification of the accumulation of anti-IgG on sensor surface was demonstrated. Additionally, for biological relevance interactions between cancer biomarker EGFr and EGF was also studied and the preliminary results are presented. The potential applications of the label free technique include point-of-care screening of cancer biomarkers and tumor cell adhesion analysis.

Gold nanoparticles have been shown to possess x-ray attenuation higher than that of clinically used iodine based contrast agents. Additionally, gold nanoparticles have also been used in enhancing the Raman signal through a phenomenon called Surface Enhanced Raman Scattering (SERS). The development of Gold nanoparticle based probes (nanotags) for combined SERS and x-ray computed tomography (CT) are reported. These nanotags comprise quasi-spherical gold nanoparticles encoded using a Raman active molecule and encapsulated by a monolayer of polyethylene glycol with carboxylate functional groups for bioconjugation. The nanotags, made from ~65 nm gold nanoparticles, display large SERS cross-section and x-ray absorption which are used for dual modality imaging of tumor cells in of an orthotropic prostate cancer tumor animal model.

Multi-color gold-nanoparticle-based tags (nanotags) are synthesized for combined surface-enhanced Raman spectroscopy (SERS) and x-ray computed tomography (CT). The nanotags are synthesized with quasi-spherical gold nanoparticles encoded with a reporter dye (color), each with a unique Raman spectrum. A library of nanotags with six different colors were synthesized for a range of gold nanoparticle sizes and an optimum size has been established to yield the largest SERS intensity and x-ray attenuation that is higher than the iodinated CT contrast agents used in clinics. Proof-of-principle in vivo imaging results with nanotags are presented that, for the first time, demonstrates the combined in vivo dual modality imaging capability of SERS and CT with a single nanoparticle probe.

We report the synthesis and characterization of multi-color gold nanoparticle based tags (Raman Nanotags) for molecular imaging. The multi-color Raman Nanotags are PEGylated gold nanoparticles (AuNPs) encapsulating a Raman reporter dyes which can be functionalized with any ligand of interest for targeted molecular imaging. The Raman Nanotags synthesized with 65 nm gold nanoparticles exhibit the largest surface enhanced Raman scattering (SERS) signal. Results are presented quantify the measured SERS signal, dynamic range, reproducibility, and stability of Raman Nanotags. In vitro cell culture experiments for targeted biomarker detection using functionalized Raman Nanotags are also presented.

A simple yet highly sensitive implementation of an interferometric technique for a label-free molecular biosensing application is described. The intereferometric detection method is based on the phase-sensitive detection of spectral interference fringes. The change in optical path length due to binding of biomolecules on functionalized optically clear substrates can be quantified by detecting the change in the phase of the spectral fringes. The common path interferometeric design permits measurement of sub-monolayer binding of biomolecules to the sensor surfaces.

A simple yet highly sensitive implementation of an interferometric technique for a label-free molecular biosensing application is described. The intereferometric detection method is based on the phase-sensitive detection of spectral interference fringes. The change in optical path length due to binding of biomolecules on functionalized optically clear substrates can be quantified by detecting the change in the phase of the spectral fringes. The common path interferometeric design permits measurement of sub-monolayer binding of biomolecules to the sensor surfaces.

We describe a dual-channel optical low-coherence reflectometer for accurate measurement of Doppler angles in highly scattering media. Accurate fluid-flow velocity estimation requires measurement of the Doppler shift and angle. Estimated values of the Doppler angle and average fluid-flow velocity from experimental data are in good agreement with preset values.

A birefringent-fiber-based dual-channel optical low-coherence reflectometer capable of differential phase measurement is described. Phase noise owing to environmental perturbations that is common to both channels is canceled, resulting in accurate measurement of the phase difference between light backscattered from two spatially separated sites. Results are presented that demonstrate the accuracy and sensitivity of the system.

A compact fiber optic scanner for biomedical applications such as optical coherent tomography has been designed, fabricated and tested. The scanner is designed as an in vivo device and composed of an optical fiber coated with nickel-powder loaded paint for external magnetic actuation. The compactness of the imaging device makes it suitable for applications where size, precision and low power consumption is critical. We have previously demonstrated the principles utilizing magnetic actuation for the fiber scanner coated with magnetic gel. This work focused on verification and optimization of the scanner operation. The magnetic properties of the nickel particle mixed with paint were characterized using an alternating gradient magnetometer. The optical scanner is externally actuated by an electromagnet and so it does not require a voltage or current supply in the probe itself. The displacements of the scanner were recorded using a position sensitive detector. The result showed a 0.8-mm displacement under the influence of a static magnetic field of 17.6 KA/m in a fiber with a moveable length of 4.2cm. Dynamic analysis showed a displacement of 0.83mm with an input current amplitude of 41mA and a magnetic field of 2.4 KA/m. The measurements are in good agreement with the theoretical lumped-element calculations. Finite-element analysis was performed and the results agree with the theoretical and experimental results. The static and dynamic displacements of the fiber optic scanner depend on the thickness and length of the magnetic coating. Thus, scanners for different displacements and operating frequencies can be designed by varying the coating thickness and length.

We have demonstrated non-contact, sub-nanometer optical measurement of neural surface displacement associated with action potential propagation without applying exogenous chemicals or reflection coatings. Signals recorded from crayfish leg nerve using a phase-sensitive optical low coherence reflectometer show that transient neural surface displacement due to action potential propagation is approximately 1 nm in amplitude and 1 ms in duration. Measured optical signals are coincident with electrical action potential arrival to the optical measurement site. Recent experiments indicate signals with similar amplitude and duration are observed in response to repetitive fast stimulation (200 stimuli/s).

We describe a method for en face phase-contrast imaging of cells with a fiber-based differential phase-contrast optical coherence microscopy system. Recorded en face images are quantitative phase-contrast maps of cells due to spatial variation of the refractive index and (or) thickness of various cellular components. Quantitative phase-contrast images of human epithelial cheek cells obtained with the fiber-based differential phase-contrast optical coherence microscopy system are presented.

We describe a methodology to record spatial variation of refractive index of porcine renal artery using differential phase optical coherence tomography (DP-OCT). DP-OCT provides a quantitative measure of thin specimen phase retardation and refractive index with phase resolution of 5 nm and lateral resolution of 3 mm. DP-OCT instrumentation is an all-fiber, dual channel Michelson interferometer constructed using polarization maintaining fiber. Two orthogonal polarization modes of light are spatially separated using a Wollaston prism and directed into separate photoreceivers. Because phase noise in the environment is equally present in both channels, computation of phase difference between the two signal channels is attributed exclusively to variation in the specimen's composite refractive index. Porcine renal artery is freshly harvested from a local slaughter house. The lumen is cut open and sliced at 5 mm thickness. Microscopic slide for the tissue section is processed by standard histology method with mounting media. Two dimensional en face dual-channel phase images are taken over 150 mm x 200 mm region on the microscopic slide and the images are reconstructed by plotting relative phase variation as the OCT beam is moved across the artery cross section.

Optical coherence tomography (OCT) is a novel technique that has recently emerged as a promising biomedical imaging modality in ophthalmology, cardiology, oncology and a number of other medical disciplines. Basic contrast mechanism for OCT is gradient in refractive index of tissue constituents. Additional contrast mechanisms include Doppler, polarization and phase-sensitive imaging modalities. Applications of metrology include high-resolution spectrum analyzers for spectral domain OCT instrumentation development and characterization of refractive index of tissue constituents.

Tissue response to thermal, electrical, or chemical stimuli are important in the health and survival of tissue. We report experimental results to assess tissue response to various stimuli using a low coherence differential phase interferometer.The optical system utilized to measure tissue response is a novel fiber-based phase sensitive optical low coherence reflectometer (PS-OLCR). Inasmuch as the PS-OLCR works with back-reflected light, noninvasive sensing of tissue response to stimuli is possible. In addition to high lateral (approximately 10 microm) and longitudinal (approximately 10 microm) resolution, PS-OLCR can measure sub-wavelength changes in optical path-length (Angstrom/nanometer range) by extracting the phase difference between interference fringes in two channels corresponding to orthogonal polarization modes.When light spatially splits into two polarization states, precise analysis of surface topography or tissue surface response such as swelling or collapse are possible. Time resolved measurements of nanometer-scale path length changes in response to electrical and thermal stimuli are demonstrated using longitudinally delayed polarization channels.Since PS-OLCR is a useful tool to detect ultra-small path length changes, the system has potential to aid scientists in investigating important phenomena in biomaterials and developing useful diagnostic and therapeutic imaging modalities. Applications include tissue surface profilometry, measurement of tissue, and cell response to various stimuli, high-resolution intensity and phase imaging.

A fiber based dual channel phase-sensitive optical low coherence tomography system is proposed for birefringence imaging neural tissues. The system measures relative optical path length differences between the orthogonal modes of the polarization-maintaining fiber in highly scattering media with high lateral and longitudinal spatial resolution. Because of the ability of detecting ultra-small changes in refractive index, system has uses in many bio-medical optical imaging applications such as monitoring cell and tissue dynamics, imaging of phase objects, and functional studies.

We describe a phase sensitive optical low coherence reflectometer using polarization maintaining optical fiber with high lateral, longitudinal and phase resolution for biomedical applications. The system utilizes three sample path configurations (i.e. single beam, lateral beams, and longitudinally delayed channels) and allows investigation of various topics in biomedical research. Applications include measurement of birefringence change, intensity and phase imaging, refractive index measurements, surface analysis, and measurement of solute concentrations.

We have developed a low-coherence optical sensor for detection of laser-induced thermoelastic deformations in biological materials. The presented optical sensor utilizes a birefringent fiber-based dual channel low-coherence Michelson interferometer capable of differential phase measurements. We demonstrate that the low-coherence sensor can be used for spatially-resolved measurements of laser-induced thermoelastic deformations in biological materials with high axial resolution. Experimental studies were carried out using gelatin-based tissue phantoms.

Imaging the birefringence characteristics of the retinal nerve fiber layer (RNFL) may aid in glaucoma diagnosis. Polarization Sensitive Optical Coherence Tomography (PSOCT) was used to record in vivo high resolution images of the RNFL in two cynomologous monkeys. Depth variation in the Stokes vector of reflected light was used to calculate the optical phase retardation per unit depth (OPR/UD) as a function of retinal position. OPR/UD decreased from 32/spl deg//100 /spl mu/m near the optic nerve to 5/100 /spl mu/m at a location 600 /spl mu/m nasal to the optic nerve. Variation of OPR/UD in the RNFL with retinal position demonstrates a chance in birefringence for different densities of ganglion cell axons. PSOCT may be useful for noninvasive determination of RNFL thickness and fiber density.

The results presented here show that field control of second‐harmonic generation (SHG) can be substantially enhanced in compositionally asymmetrical wells. In an optimized step well the magnitude of second order susceptibility for moderate field (±80 kV/cm) is 2 orders of magnitude larger than that in the square well with the same applied field. Numerical results are presented for SHG at a CO2 wavelength for a square, graded, and step well.

We present a fiber optic spectral polarimeter, an instrument that allows measurement of the spectrally resolved Stokes parameters. The instrument consists of a pair of polarization maintaining (PM) fibers spliced at 45 degree(s) with respect to each other in line with a polarizer module whose transmission axis is aligned to the fast axis of the first PM fiber. The generated spectrum is composed of three quasi- cosinusoidal components, which carry information required to reconstruct the spectrally resolved Stokes parameters of incident light. Fourier transform of the measured spectrum provides the significant parameters for determination of the spectrally resolved Stokes parameters of light. The performance of the fiber optic spectral polarimeter is demonstrated by the computer simulation and experiments with input light of known polarization states. A unique characteristic of this instrument is that the spectrally resolved polarization state of incident light can be determined from one spectral measurement without any mechanical movement. Moreover, the instrument can be incorporated into a tomographic imaging system such as a conventional microscopy system. Polarization based microscopy combined with a fiber optic spectral polarimeter will provide measurements that allow better understanding of depolarization process of light passing through biological materials for diagnostic imaging.© (2002) COPYRIGHT SPIE--The International Society for Optical Engineering. Downloading of the abstract is permitted for personal use only.