This PDF file contains the front matter associated with SPIE Proceedings Volume 8580, including the Title Page, Copyright information, Table of Contents, and the Conference Committee listing.

2D/3D spatial distribution of superficial blood vessels in human skin in vivo was conducted by double correlation analysis of the swept source Optical Coherence Tomography (OCT) images. An adaptive Wiener filtering technique has been employed to remove background noise and increase the overall quality of the OCT images acquired experimentally. Correlation Mapping and Fourier domain correlation approaches have been subsequently applied to enhance spatial resolution of images of vascular network in human skin. The analysis of images performed on Graphics Processing Units (GPUs) utilizing the recently developed Compute Unified Device Architecture (CUDA) framework.

The retina/choroid structure is an example of a complex biological target featuring highly perfused tissues and vessel flows both near the surface and at some depth. Laser speckle imaging can be used to image blood flows but static scattering paths present a problem for extracting quantifiable data. The speckle contrast is artificially increased by any residual specular reflection and light paths where no moving scatterers are encountered. Here we present results from phantom experiments demonstrating that the static and dynamic contributions to laser speckle contrast can be separated when camera exposures of varying duration are used. The stationary contrast parameter follows the thickness and strength of the overlying scatterer while the dynamic proportion of the scatter resulting from vessel flows and Brownian motion is unchanged. The importance of separating the two scatter components is illustrated by in vivo measurements from a scarred human retina, where the effect of the un-perfused scar tissue can be decoupled from the dynamic speckle from the intact tissue beneath it.

How does speckle contrast K, measured at camera exposures T around 10 ms, give us information about temporal autocorrelation of the speckle pattern with time constants τ < 1 ms, corresponding to Doppler shifts in the KHz range? We explore the implications of this question and show that for any particular assumed temporal speckle autocorrelation function, K measured at T >> τ accurately measures τ, but that K measurements at T < τ are required in order to determine the actual shape of the autocorrelation function. Determining the shape of the autocorrelation function is important if we wish to distinguish between different types of flow or movement in tissue, for example distinguishing Brownian motion or the randomly-oriented flows in capillary networks from more ordered flow in resolvable vessels.

In this study, we tested the hypothesis that a microbubble containing photosensitizer when activated with light would enable comprehensive disinfection of bacterial biofilms in infected root dentin by antimicrobial photodynamic therapy (APDT). Experiments were conducted in two stages. In the stage-1, microbubble containing photosensitizing formulation was tested for its photochemical properties. In the stage-2, the efficacy of microbubble containing photosensitizing formulation was tested on in vitro infected root canal model, developed with monospecies biofilm models of Enterococcus faecalis on root dentin substrate. The findings from this study showed that the microbubble containing photosensitizing formulation was overall the most effective formulation for photooxidation, generation of singlet oxygen, and in disinfecting the biofilm bacteria in the infected root canal model. This modified photosensitizing formulation will have potential advantages in eliminating bacterial biofilms from infected root dentin.

There is a need in tissue engineering to monitor cell growth and health within 3D constructs non-invasively and in a label-free manner. We have previously shown that optical coherence phase microscopy was sensitive enough to monitor intracellular motion. Here we demonstrate that intracellular motility can be used as an endogeneous contrast agent to image cells in various 3D engineered tissue architectures. Phase and intensity-based reconstruction algorithms are compared. In this study, we used an optical coherence phase microscope set up in a common path configuration, developed around a Callisto OCT engine (Thorlbas) centred at 930nm and an inverted microscope with a custom scanning head. Intensity data were used to perform in-depth microstructural imaging. In addition, phase fluctuations were measured by collecting several successive B scans at the same location, and the first time derivative of the phase, i.e. time fluctuations, was analysed over the acquisition time interval to map the motility. Alternative intensity-based Doppler variance algorithms were also investigated. Two distinct scaffold systems seeded with adult stem cells; algimatrix (Invitrogen) and custom microfabricated poly(D,L-lactic-co-glycolic acid) fibrous scaffolds, as well as cell pellets were imaged. We showed that optical phase fluctuations resulting from intracellular motility can be used as an endogenous source of contrast for optical coherence phase microscopy enabling the distinction of viable cells from the surrounding scaffold.

Polyamidoamine (PAMAM) dendrimers have been topically applied to the skin and utilised as a permeation enhancer for a range of therapeutic compounds. However, very little is known about the mechanism of enhancement. This study used optical coherence tomography (OCT) to investigate the influence of PAMAM dendrimers to alter surface refractive index (RI) in excised porcine skin. It is revealed that PAMAM dendrimers caused a sporadic disruption and disappearance of the white hyper-reflective band on the skin surface using OCT. Following the decontamination of the treated skin specimens, the entrance signal, resulting in the polarised light reflecting off the keratin of the upper skin strata, returned to normal. Further, PAMAM-induced changes in skin RI was benchmarked against glycerol, a known permeation enhancer and skin clearing agent. Changes in RI with PAMAM were only observed on the skin surface, suggesting that the dendrimer only modulates the outer layers of the stratum corneum. This is substantially different to the observed effect of glycerol, which permeated more deeply into the skin. The non-invasive and non-destructive OCT imaging technique may provide a convenient tool to investigate the mechanism of permeation enhancement and transdermal drug delivery.

As one of the important components of optical microscopes, the condenser has a considerable impact on system performance, especially on the depth of field (DOF). DOF is a critical technical feature in cytogenetic imaging that may affect the efficiency and accuracy of clinical diagnosis. The purpose of this study is to investigate the influence of microscopic condenser on DOF using a prototype of transmitted optical microscope, based on objective and subjective evaluations. After the description of the relationship between condenser and objective lens and the theoretical analysis of the condenser impact on system numerical aperture and DOF, a standard resolution pattern and several cytogenetic samples are adopted to assess the condenser impact on DOF, respectively. The experimental results of these objective and subjective evaluations are in agreement with the theoretical analysis and show that, under the specific intermediate range of condenser numerical aperture ( NAcond ), the DOF value decreases with the increase of NAcond . Although the above qualitative results are obtained under the experimental conditions with a specific prototype system, the methods presented in this preliminary investigation could offer useful guidelines for optimizing operational parameters in cytogenetic imaging.

We describe a multiple reference OCT (MR-OCT) system which is radically different from existing optical coherence tomography (OCT) systems. Complete scans of a target from surface to depth are accomplished by simultaneously acquiring the scan in multiple segments using a virtually solid state design that is inherently miniature, robust and low cost; in short, ideal for use in applications characterized by high volumes, difficult operating environments and constrained acquisition and operating budgets.

We describe a novel application of correlation mapping optical coherence tomography (cmOCT) for sub-surface fingerprint biometric identification. Fingerprint biometrics including automated fingerprint identification systems, are commonly used to recognise the fingerprint, since they constitute simple, effective and valuable physical evidence. Spoofing of biometric fingerprint devices can be easily done because of the limited information obtained from the surface topography. In order to overcome this limitation a potentially more secure source of information is required for biometric identification applications. In this study, we retrieve the microcirculation map of the subsurface fingertip by use of the cmOCT technique. To increase probing depth of the sub surface microcirculation, an optical clearing agent composed of 75% glycerol in aqueous solution was applied topically and kept in contact for 15 min. OCT intensity images were acquired from commercial research grade swept source OCT system (model OCT1300SS, Thorlabs Inc. USA). A 3D OCT scan of the fingertip was acquired over an area of 5x5 mm using 1024x1024 A-scans in approximately 70 s. The resulting volume was then processed using the cmOCT technique with a 7x7 kernel to provide a microcirculation map. We believe these results will demonstrate an enhanced security level over artificial fingertips. To the best of our knowledge, this is the first demonstration of imaging microcirculation map of the subsurface fingertip.

The two highest principal components of fluorescence spectra in visible region obtained, using Xenon lamp as an excitation source of normal and dysplastic human cervical tissues are analyzed using scatter plots and probability density functions. These yield significant differences between the tissue types.

Chemical agents with high refractive index, hyperosmotic, and biocompatibility are introduced into tissue, which will reduce the scattering of tissue, and enhance the penetration of light in tissue. Diffuse reflectance, as a common method, has been applied to assess optical clearing of skin in vivo, but the scattering characteristic during the in-vivo optical clearing process has not been valuated quantitatively. In this work, a diffuse reflectance spectroscopy, based on a lookuptable (LUT) based inverse model, is applied to calculate the reduced scattering coefficient and absorption coefficient of skin. Optical clearing agents (OCAs) were topically treated on mouse skin in vivo. The diffuse reflectance during optical clearing was recorded, and the optical properties can be extracted by the reflectance spectroscopy. The results show that the diffuse reflectance spectra and the reduced scattering coefficient are decreased obviously, whilst the absorption coefficient is increased after the application of OCAs. This study provides evident directly for explore the mechanisms of optical clearing of skin in vivo.

We present a computational method for the analysis of optical coherence tomography (OCT) images to detect soft tissue sarcomas. The method combines the quantitative analysis of two aspects of information from the intensity A-lines of OCT images; one is the slope of the intensity A-line with dB unit, which is determined by the optical attenuation characteristics of tissue; the other is the standard deviation (SD) of the slope-removed intensity A-line, which is dependent on the tissue structural features. The method is tested with pilot experiments on ex vivo tissue samples of human fat, muscle, well differentiated liposarcoma (WDLS) and leiomyosarcoma. Our results demonstrate the feasibility of this quantitative method in the differentiation of soft tissue sarcomas from normal tissues. This study indicates that OCT can be a potential computer-aided means of automatically and accurately identifying resection margins of soft tissues sarcomas during surgical treatment.

The vascular system of Zebrafish embryos is studied by means of Fluorescence Correlation and Image Correlation Spectroscopy. The long term project addresses biologically relevant issues concerning vasculogenesis and cardiogenesis and in particular mechanical interaction between blood flow and endothelial cells. To this purpose we use Zebrafish as a model system since the transparency of its embryos facilitates morphological observation of internal organs in-vivo. The correlation analysis provides quantitative characterization of fluxes in blood vessels in vivo. We have pursued and compared two complementary routes. In a first one we developed a two-spots two-photon setup in which the spots are spaced at adjustable micron-size distances (1-40 μm) along a vessel and the endogenous (autofluorescence) or exogenous (dsRed transgenic erythrocytes) signal is captured with an EM-CCD and cross-correlated. In this way we are able to follow the morphology of the Zebrafish embryo, simultaneously measure the heart pulsation, the velocity of red cells and of small plasma proteins. These data are compared to those obtained by image correlations on Zebrafish vessels. The two methods allows to characterize the motion of plasma fluids and erythrocytes in healthy Zebrafish embryos to be compared in the future to pathogenic ones.

Previously we demonstrated the utility of optical fluorometry to evaluate lung tissue mitochondrial redox state in isolated perfused rats lungs under various chemically-induced respiratory states. The objective of this study was to evaluate the effect of acute ischemia on lung tissue mitochondrial redox state in vivo using optical fluorometry. Under ischemic conditions, insufficient oxygen supply to the mitochondrial chain should reduce the mitochondrial redox state calculated from the ratio of the auto-fluorescent mitochondrial metabolic coenzymes NADH (Nicotinamide Adenine Dinucleotide) and FAD (Flavoprotein Adenine Dinucleotide). The chest of anesthetized, and mechanically ventilated Sprague-Dawley rat was opened to induce acute ischemia by clamping the left hilum to block both blood flow and ventilation to one lung for approximately 10 minutes. NADH and FAD fluorescent signals were recorded continuously in a dark room via a fluorometer probe placed on the pleural surface of the left lung. Acute ischemia caused a decrease in FAD and an increase in NADH, which resulted in an increase in the mitochondrial redox ratio (RR=NADH/FAD). Restoration of blood flow and ventilation by unclamping the left hilum returned the RR back to its baseline. These results (increase in RR under ischemia) show promise for the fluorometer to be used in a clinical setting for evaluating the effect of pulmonary ischemia-reperfusion on lung tissue mitochondrial redox state in real time.

We present a full range spectral-domain correlation mapping optical coherence tomography (cm-OCT) method to obtain complex conjugate free, full-range correlation mapping angiography. The mirror image elimination is based on linear phase modulation of B-frames by introducing a slight off-set of the probe beam with respect to the lateral scanning fast mirror’s pivot axis. An algorithm that exploits Hilbert transform to obtain complex-conjugate free image in conjunction with the cm-OCT algorithm is used to obtain full-range imaging of microcirculation within tissue beds in vivo. The proposed system is based on a high speed spectrometer at 92kHz with a modified scanning protocol to achieve higher acquisition speed to render cmOCT images with high-speed and wide scan range. The estimated sensitivity of the system was around 105dB near the zero-delay line with ~13dB roll-off from 0.5mm to 3mm imaging-depth position. The estimated axial and lateral resolutions are 12 μm and 25 μm, respectively. A direct consequence of this complex conjugate artifact elimination is the enhanced flow imaging sensitivity for deep tissue imaging application by doubling the imaging depth. In turn, this also provides additional flexibility to explore the most sensible measurement range near the zero-delay line.

Continuous monitoring of cerebral blood oxygenation is critically important for the management of many lifethreatening conditions. Non-invasive monitoring of cerebral blood oxygenation with a photoacoustic technique offers advantages over current invasive and non-invasive methods. We introduce a Monte Carlo XYZ-PA to model the energy deposition in 3D and the time-resolved pressures and velocity potential based on the energy absorbed by the biological tissue. This paper outlines the benefits of using Monte Carlo XYZ-PA for optimization of photoacoustic measurement and imaging. To the best of our knowledge this is the first fully integrated tool for photoacoustic modelling.

Skipping various numbers of A-lines is effective to obtain multi-range velocimetry using Doppler optical coherence tomography (DOCT). High correlation between A-lines is a fundamental prerequisite for DOCT processing. Therefore, high oversampling is normally necessary, especially when skipping A-lines. That requires quite a long time of imaging, which might not be acceptable on some occasions. Step-scanning protocol, which captures repeated A-scans, has been employed for multi-range DOCT previously. We develop it by waiting for the scanner to stabilize, but not capturing continuously. In this way the cross-correlation of step scanning maintains almost constant for all the captured A-lines, and is higher than that of conventional linear scanning with similar imaging time. Due to the limited numbers of A-lines at each lateral position, we choose previously proposed high-pass filter in ultrahigh sensitive optical microangiography (OMAG) to enhance flow sensitivity. Doppler processing is implemented after the filter, both of which utilize A-line skipping to achieve variable velocity ranges. The obtained Doppler signal in blood flow is encircled by much noise in non-flow area. This is because that the static components are rejected by the filter, leaving random phase noise. The phase variance, which is a flow indicator, is employed to generate a binary mask to extract the Doppler signal out of noise. The technique is demonstrated in bi-directional cross-section and maximum projection en face images of middle cerebral artery (MCA) occluded mouse model. The vasculature responses from artery down to capillary during baseline, occlusion and reperfusion are illustrated. In some arteries that branch from MCA the flow is reversed but not simply reduced or vanished.

Previous studies have preliminarily validated the floating reference method and shown that it has the potential to improve the accuracy of non-invasive blood glucose sensing by Near-Infrared Spectroscopy. In order to make this method practical, it is necessary to precisely verify and measure the existence and variation features of the positional floating reference point. In this talk, a device which can precisely verify and measure the positional floating reference point is built. Since the light intensity of diffuse reflectance from the tested sample is very weak, a multipath detecting fibers system was built to improve signal-to-noise ratio. In this system, the fibers encircle the light source fiber which is regarded as the reference center of detecting fibers while they are moving. In addition, the position of each fiber is accurately controlled by manual translation stage to keep all detecting fibers always in the same radius around light source fiber. This ensures that received signal is coming from the same radial distance of light source. The variation of signal-to-noise ratio along with the different radial distance was investigated based on experiments. Results show that the application of this device could improve signal-to-noise ratio, and provide a new experimental method for the further study of positional floating reference point.

Non-invasive measurement of human body composition by Near-Infrared(NIR) spectroscopy is one of hot research topics in Biomedical Photonics field. However, the practical application of this technique is not available, yet because of the uncertainty of measurement conditions during in vivo measurement. The uncertainty would greatly influence the acquisition of spectral signal which reflects different concentrations of the constituent of human body. The location of optical sensor has been proved to be associated with the diffused reflection light. In this talk, a computer vision localization method was proposed, which is based on image matching to localize optical sensor and further the impact of optical sensor position and orientation variance can be decreased. Experiment results showed that the uncertainty of measurement could be reduced significantly.

Previous studies have shown the limitations of taking OGTT (Oral Glucose Tolerance Test) as the glucose adjustment protocol for non-invasive blood glucose sensing. Previous studies built a mathematical model of glucose metabolism system-IMM (the Integrated Minimal Model) to probe other available adjustment methods. In this talk, a further study would be focused on more detailed combination options of different glucose input types for glucose adjustment projects in non-invasive blood glucose sensing. And predictive models of blood glucose concentration have been established by means of partial least squares (PLS) method, which could be used to evaluate the quality of different glucose adjustment options. Results of PLS modeling suggested that predictive models under combined glucose input types, compared with OGTT, show a great enhancement in the stability. This would finally improve the precision of non-invasive blood glucose sensing.

Near-infrared spectroscopy(NIRs) is an ideal measurement method for noninvasive blood glucose sensing. In that measuring process, the light propagation path in skin tissue would affect the final near-infrared spectrum greatly. In this talk, the Monte Carlo simulation has been conducted to investigate the effect of dermal thickness variances on blood glucose sensing results at earlobe site. Results demonstrate that the floating reference point exists in the finite five-layered thickness skin model, and the variation of floating reference point under the dermal thickness change is simulated. It is indicated that in the dermal thickness increasing process, there is a rising trend of the floating reference point at each wavelength. It will lay a solid foundation for the further design of an advanced blood glucose detection probe to facilitate the application of NIRs to noninvasive blood glucose sensing eventually.

The problem of automatic recognition of specific oscillatory patterns on electroencephalograms (EEG) is addressed using the continuous wavelet-transform (CWT). A possibility of improving the quality of recognition by optimizing the choice of CWT parameters is discussed. An adaptive approach is proposed to identify sleep spindles (SS) and spike wave discharges (SWD) that assumes automatic selection of CWT-parameters reflecting the most informative features of the analyzed time-frequency structures. Advantages of the proposed technique over the standard wavelet-based approaches are considered.

Spike-wave discharges are electroencephalographic hallmarks of absence epilepsy. Spike-wave discharges are known to originate from thalamo-cortical neuronal network that normally produces sleep spindle oscillations. Although both sleep spindles and spike-wave discharges are considered as thalamo-cortical oscillations, functional relationship between them is still uncertain. The present study describes temporal dynamics of spike-wave discharges and sleep spindles as determined in long-time electroencephalograms (EEG) recorded in WAG/Rij rat model of absence epilepsy. We have proposed the wavelet-based method for the automatic detection of spike-wave discharges, sleep spindles (10–15Hz) and 5–9Hz oscillations in EEG. It was found that non-linear dynamics of spike-wave discharges and sleep spindles fits well to the law of ’on-off intermittency’. Intermittency in sleep spindles and spike-wave discharges implies that (1) temporal dynamics of these oscillations are deterministic in nature, and (2) it might be controlled by a system-level mechanism responsible for circadian modulation of neuronal network activity.

Wavelets are widely used to improve the quality of images by digital filtering of noise. In this work, application of wavelet-filtering to OCT-images is considered. The problem of appropriate selection of the wavelet-basis is discussed and analysis of cerebral arteries in newborn rats is performed.

Congenital Heart Disease (CHD) is the most common congenital malformation in newborns in the US. Although knowledge of CHD is limited, altered hemodynamic conditions are suspected as the factor that stimulates cardiovascular cell response, resulting in the heart morphology remodeling that ultimately causes CHDs. Therefore, one of recent efforts in CHD study is to develop high-speed imaging tools to correlate the rapidly changing hemodynamic condition and the morphological adaptations of an embryonic heart in vivo. We have developed a high-speed streak mode OCT that works at the center wavelength of 830 nm and is capable of providing images (292x220 μm²) of the outflow tract of an embryonic chick heart at the rate of 1000 Hz. The modality can provide a voxel resolution in the range of 10 μm³, and the spectral resolution allows a depth range of 1.63 mm. In the study reported here, each of the 4D images of an outflow tract was recorded for 2 seconds. The recording was conducted every 2 hours (HH17 to HH18), 3 hours (HH14 to HH17), and 4 hours (HH18 to HH19). Because of the fast scan speed, there is no need for postacquisition processing such as use of gating techniques to provide a fine 3D structure. In addition, more details of the outflow tract are preserved in the recorded images. The 4D images can be used in the future to determine the role of blood flow in CHD development.

A mathematical model and numerical simulation of nonstationary temperature field within tissues and cells locally heated under pulse laser irradiation mediated by plasmonic nanoparticles are presented. The essential temperature alterations during laser pulse and time interval between pulses was found. Temperature spatial-temporal profile depends on a number of parameters including optical and thermophysical properties of cell membrane and nanoparticles, laser wavelength, energy, pulse duration and repetition rate. On the basis of introduced parameters a quantitative evaluation of thermal effects was done. The possibilities of usage of these parameters to study their correlation with irreversible photothermal effects, including a change in the permeability of cell membrane during optoporation, are discussed.