This PDF file contains the front matter associated with SPIE Proceedings Volume 8222, including the Title Page, Copyright information, Table of Contents, Introduction, the Report on the Panel Discussion, and the Conference Committee listing.

A variety of laser Doppler and laser speckle contrast systems have been constructed by various groups and companies, for commercial sale and for research. All rely on the same physical phenomenon - the dynamic laser speckle pattern generated by illuminating tissue with coherent light - but differ in details of system design, operation and analysis. We present a comparison between measurements made with three systems: a multi-exposure laser speckle contrast system built at Industrial Research Ltd, a commercial single-exposure laser speckle contrast system developed by Perimed AB (PSI NR) and the full-field laser Doppler camera built by the University of Twente (TOPCam). We compare the response to changing flows of all three systems. The systems are found to produce similar results for a variety of in-vivo and in-vitro measurements. Multi-exposure speckle contrast shows some advantages in information gained and insensitivity to static speckle, at the cost of increased complexity and measurement time.

Measurements of flow in retinal vessels is presented and compared with in vitro measurements on whole blood in capillaries ranging from 75 to 200μm diameter. The viewing angle of the capillaries and their range of size allows size-dependent effects to be investigated when estimating flow within actual vessels. Retinal measurements show a pulse effect. When this is removed by synchronisation, multi-exposure measurements show different spectral signature from single speed scatterers. Multi-exposure measurements of the retina demonstrate a varying contribution of stationary scatter across the field. Unlike scattering in dermal tissue, photons in retinal vessels must return by multiple scatter from moving Red Blood Cells, whose motion is directed. Speckle estimates of flow in retinal vessels are therefore possible.

Blood protein molecules could be embedded in porous glassy substrate with 10-nm pores. The embedding principle is based on blood cell dehydration with the destruction of the cell membrane, and reconstitution and centrifuge could yield a suitable solution for doping into a porous glassy medium. The doped glassy substrate speckle pattern under laser illumination could be used to characterize the protein size distribution. Calibration with known protein embedded samples would result in an optical procedure for the characterization of a blood sample. Samples embedded with larger kilo-Dalton protein molecule show more variation in the speckle patterns, consistent with protein folding interaction inside a pore cavity. A regression model has been used to correlate the protein molecule sizes with speckle sizes. The use of diffusion mean free path information to study protein folding in the embedding process is briefly discussed.

We present a dual-wavelength endoscopic laser speckle contrast system including illumination with polarization maintaining fibres and imaging using a leached fibre image guide. This system has a frame rate of 10 Hz and can rapidly monitor changes in blood flow in vivo, including due to the heart beat, using the contrast values of the speckle images recorded with 1 ms exposure time. In addition the mean intensities can record the respiration period and can indicate changes in tissue oxygenation. This system was tested during an occlusion to a human finger and is being applied in endoscopy.

Atherosclerosis is the most common underlying cause of vascular disease, occurring in multiple arterial beds including the carotid, coronary, and femoral arteries. Atherosclerosis is an inflammatory process occurring in arterial tissue, involving the subintimal accumulation of low-density lipoproteins (LDL). Little is known about the rates at which these accumulations occur. Measurements of the permeability rate of LDL, and other lipoproteins such as high-density lipoprotein (HDL) and very low-density lipoprotein (VLDL), could help gain a better understanding of the mechanisms involved in the development of atherosclerotic lesions. The permeation of VLDL, LDL, HDL, and glucose was monitored and quantified in normal and diseased human carotid endarterectomy tissues at 20°C and 37°C using optical coherence tomography (OCT). The rates for LDL permeation through normal tissue at 20°C was (3.16 ± 0.37) × 10^-5 cm/sec and at 37°C was (4.77 ± 0.48) × 10^-5 cm/sec, significantly greater (p<0.05) than the rates for diseased tissue at these temperatures ((1.97 ± 0.34) × 10^-5 cm/sec and (2.01 ± 0.23) × 10^-5 cm/sec, respectively). The observed results support previous suggestions of an enhanced transport mechanism specific to LDL. This study effectively uses optical coherence tomography to measure the rates of permeation of vascular tissue by the range of naturally occurring lipoproteins.

In this work, we report a wavelet based multi-fractal study of images of dysplastic and neoplastic HE- stained human cervical tissues captured in the transmission mode when illuminated by a laser light (He-Ne 632.8nm laser). It is well known that the morphological changes occurring during the progression of diseases like cancer manifest in their optical properties which can be probed for differentiating the various stages of cancer. Here, we use the multi-resolution properties of the wavelet transform to analyze the optical changes. For this, we have used a novel laser imagery technique which provides us with a composite image of the absorption by the different cellular organelles. As the disease progresses, due to the growth of new cells, the ratio of the organelle to cellular volume changes manifesting in the laser imagery of such tissues. In order to develop a metric that can quantify the changes in such systems, we make use of the wavelet-based fluctuation analysis. The changing self- similarity during disease progression can be well characterized by the Hurst exponent and the scaling exponent. Due to the use of the Daubechies' family of wavelet kernels, we can extract polynomial trends of different orders, which help us characterize the underlying processes effectively. In this study, we observe that the Hurst exponent decreases as the cancer progresses. This measure could be relatively used to differentiate between different stages of cancer which could lead to the development of a novel non-invasive method for cancer detection and characterization.

In the cell cycle, mitosis is the most dramatic phase, especially in Telophase and Cytokinesis. For single cells and cell monolayer, there are precise microscopic studies of mitosis, while for 3-D tissue such as tumor spheroids the light signal is obscured by the high background of diffusely scattered light. Therefore, the mitosis phase cannot be detected deep inside 3-D tissue using conventional microscopic techniques. In this work, we detect mitosis in living tissue using Tissue Dynamic Imaging (TDI). We trace depth-gated dynamic speckles from a tumor spheroid (up to 1mm in diameter) using coherence-gated digital holography imaging. Frequency-versus-time spectrograms depend on specific types of perturbation such as cell shape change, membrane undulation and cell organelles movements. By using these spectral responses as functional finger prints, we can identify mitosis events from different voxels at a specified depth inside tumor spheroids. By performing B-scans of the tumor spheroid, we generate 3-D mitosis maps (or movies) for the entire tumor spheroids. We show that for healthy tumor spheroids, the mitosis events only happen within the proliferating shell. We also compare results when anti-cancer drugs are applied to arrest, release and synchronize mitosis. This shows the application of TDI for drug screening. The technique can identify and monitor complex motilities inside 3-D tissue with a strong potential for drug diagnosis and developmental biology studies.

The quantification of (blood) flow velocity within the vasculature has potent diagnostic and prognostic potential. Assessment of flow irregularities in the form of increased permeability (micro haemorrhaging), the presence of avascular areas, or conversely the presence of vessels with enlarged or increased tortuosity in the acral regions of the body may provide a means of non-invasive in vivo assessment. If assessment of dermal flow dynamics were performed in a routine manner, the existence and prevalence of ailments such as diabetes mellitus, psoriatic arthritis and Raynaud's condition may be confirmed prior to clinical suspicion. This may prove advantageous in cases wherein the efficacy of a prescribed treatment is dictated by a prompt diagnosis and to alleviate patient discomfort through early detection. Optical Coherence Tomography (OCT) is an imaging modality which utilises the principle of optical interferometry to distinguish between spatial changes in refractive index within the vasculature and thus formulate a multi-dimensional representation of the structure of the epi- and dermal skin layers. The use of the Doppler functionality has been the predominant force for the quantification of moving particles within media, elucidated via estimation of the phase shift in OCT A-scans. However, the theoretical formulation for the assessment of these phase shifts dictates that the angle between the incident light source and the vessel under question be known a priori; this may be achieved via excisional biopsy of the tissue segment in question, but is counter to the non-invasive premise of the OCT technique. To address the issue of angular dependence, an alternate means of estimating absolute flow velocity is presented. The design and development of a dual-beam (db) system incorporating an optical switch mechanism for signal discrimination of two spatially disparate points enabling quasi-simultaneous multiple specimen scanning is described. A crosscorrelation (c-c) of interference fluctuations between these positions is performed computationally, yielding a transit time for particle flow. This paper summarises the findings of the c-c db-Sd-OCT technique for absolute velocity estimation within capillary phantoms of various sizes using Intralipid^TM solution to emulate red blood corpuscles (RBCs) and related blood constituents, driven by a calibrated syringe flow pump. The findings of the preliminary experimentation reveal the technique to be capable of estimating absolute velocity values with a maximum error difference of 0.077 mm s^-1 using Bland Altman plots. Application of this technique and rigorous testing of the c-c db-Sd-OCT method with biological samples will be the focus of future work.

An important indicator of health status is hematocrit (HCT), the fractional volume of red blood cells. As such, a noninvasive, real-time means of its measurement is highly desirable in a space flight situation. We propose to exploit ultrasound technology to provide such a capability. We introduce a specific measurement concept including the hardware and requisite processing algorithms, and discuss progress towards realization of such a measurement capability.

The formation of new microvasculature is essential for a tumor mass to grow. Vascular targeting agents (VTAs), including anti-angiogenic drugs and vascular disrupting agents, aim to either inhibit new vasculature growth or destroy existing vasculature, respectively. Because the mechanisms for anti-angiogenic drugs and vascular disrupting agents are complementary, analysis of these drugs used together is under investigation for the enhanced treatment of tumors in comparison to each treatment alone. The preclinical evaluation of the effects of VTAs on tumor growth in small animal models is vital for the development of effective drugs for clinical use. In vivo hyperspectral imaging microscopy of hemoglobin saturation has been used previously to investigate the efficacy of VTAs through analysis of tumor microvessel oxygenation after drug administration. Combining this imaging modality with first-pass fluorescence angiographic imaging can give additional important information about the vessel morphology and blood flow changes that occur after VTA treatment, thus elucidating the relationship between microvessel structure changes and oxygenation. In this study, we report the combined use of hyperspectral and first pass fluorescence angiographic imaging to examine the relationship between vessel morphology and oxygenation of human prostate cancer tumors in mice following treatment with vascular disrupting agents, OXi4503, and anti-VEGF angiogenesis inhibitor, cediranib. Imaging of the tumors is completed before treatment as well as in the days following treatment.

In this paper we examine the potential of using photoacoustic (PA) spectroscopy for the monitoring of red blood cell (RBC) aggregation phenomena. The process of RBC aggregation has been shown to occur during periods of increased plasma fibrinogen concentration and periods of decreased blood flow (leading to diminished shear forces on the aggregates). Current techniques used to monitor RBC aggregation are invasive and do not provide an accurate assessment of the aggregation process in-vivo. We present a theoretical model for investigating the potential of PA spectroscopy for detecting and characterizing the aggregation phenomenon. We show that the signal strength increases with RBC aggregation. Experimental confirmation of the theoretical predictions is provided. Our theoretical and experimental results suggest the PA spectroscopy is capable of monitoring RBC aggregation and providing important information about changes that occur during the aggregation process as it pertains to the dynamics of aggregate formation.

We analyze statistically independent temporal components in the broadband spectrum of near-infrared data acquired on the human head during breath holding. Breath holding is used to cause strong cerebral hemodynamic changes. The signals from 698 wavelengths of 25-mm long channel and 1289 wavelengths from the 10-mm channel were analyzed using an independent component analysis algorithm. We show that in spite of a very large number of channels only two components with their characteristic optical spectra remain stable across multi-subject analysis. We demonstrate a comparison of fNIRS independent components with simultaneously acquired functional MRI data.

The ability to detect changes in the structural organization of tissue has significant diagnostic value. A polarimeter has the ability to detect these changes as it can probe the tissue sub-wavelength structural organization through polarization effects. Here we present a spectral imaging polarimeter that is based on liquid crystal technology. The system is comprised of two modules, a Stokes generator and a polarimeter. Each module employs a pair of Liquid Crystal Variable Retarders (LCVRs), which are computer-controlled birefringent devices. Additionally, the polarimeter utilizes a CCD camera to image the illuminated region, thus providing spatially resolved estimates of the complete Mueller matrix for the sample. Before the system can be employed, an overall system characterization involving all four LCVRs must be performed. This characterization defines a relationship between polarimeter measurements and the incident Stokes vectors. Here we briefly describe the calibration procedure and show example Mueller matrix images of biological tissue.

Collagen is the most important component in extracellular matrix (ECM) and plays a pivotal role in individual tissue function in mammals. During ageing, collagen structure changes, which can detrimentally affect its biophysical and biomechanical properties due to an accumulation of advanced glycation end-products (AGEs). AGEs have been linked to non-enzymatic cross-linking of proteins resulting in the alteration of mechanical properties of the tissue. In this study we investigate the influence of different aged collagens on the mechanical and contractile properties of reconstituted hydrogel constructs seeded with corneal stromal fibroblasts. A non-destructive indentation technique and optical coherence tomography (OCT) are used to determine the elastic modulus and dimensional changes respectively. It is revealed that the youngest collagen constructs have a higher elastic modulus and increased contraction compared to the older collagen. These results provide new insights into the relationship between collagen molecular structures and their biomechanical properties.

Medical and security sensing applications of Terahertz (THz) imaging are currently being developed. As a result, there is a need to further investigate the effects of THz radiation on biological systems. In this study, a 94 GHz mechanically tuned Gunn Oscillator was used to irradiate Bacillus subtilis at 94 GHz. The bacteria were cultured in trypticase soy broth (TSB) and placed in polystyrene 96 well plates. The samples where irradiated during the exponential growth phase for 1, 2, and 24 hours. Both the experimental and control plates were kept at room temperature (~25°C) and were monitored for the duration of the experiment using thermocouples interfaced with a computer via Labview software. By evaluating the absorption of each well at 600nm immediately before and after irradiation, the population density within each well was assessed. Following this, the metabolic activity of each well was measured after irradiation by adding tetrazolium dye, XTT, to the wells and evaluating the absorption of each well at 490nm after 2 hours of incubation.

Currently, blood glucose concentration levels from OGTT(Oral Glucose Tolerance Test) results are used to build PLS model in noninvasive blood glucose sensing by Near-Infrared(NIR) Spectroscopy. However, the univocal dynamic change trend of blood glucose concentration based on OGTT results is not various enough to provide comprehensive data to make PLS model robust and accurate. In this talk, with the final purpose of improving the stability and accuracy of the PLS model, we introduced an integrated minimal model(IMM) of glucose metabolism system. First, by adjusting parameters, which represent different metabolism characteristics and individual differences, comparatively ideal mediation programs to different groups of people, even individuals were customized. Second, with different glucose input types(oral method, intravenous injection, or intravenous drip), we got various changes of blood glucose concentration. And by studying the adjustment methods of blood glucose concentration, we would thus customize corresponding experimental protocols of glucose adjustment to different people for noninvasive blood glucose concentration and supply comprehensive data for PLS model.

With the changing of human diet and the future of an aging society, the number of diabetic patients is growing rapidly and steadily. The major therapeutic method to that disease is monitoring the blood glucose concentration frequently to adjust the dose of the drugs and insulin. In order to avoid the painful finger prick, we choose the ear lobe as a measurement site with finger as a reference. Firstly, we compare the blood glucose concentration results of ear lobe and finger during an oral glucose tolerance test, the results showed a good correlation of the two sites. Secondly, the three-layered skin structure of finger and ear lobe has been studied by using optical coherence tomography (OCT) technique. The result shows that the thickness of each layer at ear lobe is thinner. Finally, the difference between reflectance spectra of finger and ear lobe is compared due to the diverse skin thickness. The results still show a higher absorbance value for ear lobe. In conclusion, the ear lobe is an ideal measurement site for noninvasive blood glucose sensing.

Previous study results indicated that there is a certain influence of cholesterol on non-invasive blood glucose sensing studied with NIR spectroscopy. So, this talk aims to investigate quantitative influence of cholesterol through Partial Least Squares (PLS) modeling and Unary Linear Regression (ULR) analysis respectively. PLS modeling results indicate that glucose concentration increase with the increase of cholesterol concentration. ULR analysis results indicate that there is a positive correlation between the increment of glucose and the cholesterol concentration. And the quantitative relationship has been obtained.

Since the abnormal metabolism of bilirubin could lead to diseases in the human body, especially the jaundice which is harmful to neonates. Traditional invasive measurements are difficult to be accepted by people because of pain and infection. Therefore, the real-time and non-invasive measurement of bilirubin is of great significance. However, the accuracy of currently transcutaneous bilirubinometry(TcB) is generally not high enough, and affected by many factors in the human skin, mostly by hemoglobin. In this talk, absorption spectra of hemoglobin and bilirubin have been collected and analyzed, then the Partial Least Squares (PLS) models have been built. By analyzing and comparing the Correlation and Root Mean Square Error of Prediction(RMSEP), the results show that the Correlation of bilirubin solution model is larger than that of the mixture solution added with hemoglobin, and its RMSEP value is smaller than that of mixture solution. Therefore, hemoglobin has influences on the non-invasive optical bilirubin sensing. In next step, it is necessary to investigate how to eliminate the influence.

Studying of nitric oxide (NO) dependent mechanisms of regulation of microcirculation in a stomach can provide important diagnostic markers of the development of stress-induced ulcer bleedings. In this work we use a multiscale analysis based on the discrete wavelet-transform to characterize a latent stage of illness formation in rats. A higher sensitivity of stomach vessels to the NO-level in ill rats is discussed.

Possibility of photonic crystal fiber-based glucose sensor developing is discussed. We considered the concept of the use of chirped photonic crystal fiber with big grating period as a fiber refractometer and "smart" photometric cuvette, and possibility to combine refractometric and oxidase methods for glucose determination. Influence of refractive index change of the liquid medium, determined by significant increase of containing glucose, to spectral characteristics of photonic crystal fiber was studied and the optical response to insignificant change (tenths of millimoles per liter) of glucose concentration was obtained.

The high incidence and mortality of breast cancer require an effective method for early breast diagnosis. In order to investigate the optical differences among malignant tumor, benign tumor and normal human breast tissue, a commercial spectrophotometer combined with single integrating sphere was used to measure the optical properties of different types of breast tissue in the wavelength range of 400 nm to 2200 nm in vitro. The hematoxylin and eosin staining (H&E staining) are used as the standard, and to find the find possible optical markers from the corresponding absorption or scattering spectra. This work is not only used for in vitro rapid optical diagnosis, but very helpful to develop innovative optical diagnosis of breast tumor in vivo.

Accident victims and victims of explosive devices often suffer from complex maxillofacial injuries. The lips are one of the most difficult areas of the face to reconstruct after an avulsion. Lip avulsion results in compromised facial esthetics and functions of speech and mastication. The process of reconstruction requires assessment of the vascularization of grafted ex vivo engineered tissue while it is buried underneath the skin. We describe the design and animal testing of a hand-held surgical probe based upon diffuse correlation spectroscopy to assess vascularization.

The capabilities of a pulsed photoacoustic technique for monitoring blood sedimentation and red blood cell (RBC) aggregation were tested in a cuvette in vitro. Diluted blood samples with concentrations of 40% and 60% were used. In addition, the effect of dextran (T500) was investigated with blood concentrations of 40% and 60%. Optical coherence tomography was used in parallel to visualize the sedimentation process. The results show that a 1 MHz acoustic transducer can be used to detect PA signals from blood in vitro during sedimentation, and the acoustic pulse delay is a good indicator for following the sedimentation process. Dextran greatly accelerated the sedimentation process.

Paper presents and discusses the features of laser-induced thermal dynamics of the gold nanoshells, which is associated with their relatively large size and layered structure. Unlike bulk nanoparticles the existence of a novel thermal phenomenon - hoop-shaped narrow hot zone on the nanoshell surface - is found. It is caused by spatial-temporal inhomogeneities of light field diffracted by a nanoshell and corresponding absorption of laser radiation. The numerical solution of time-dependent heat conduction equation accounting for corresponding spatially inhomogeneous distribution of heating sources is presented.

Time variation of the adipose tissue refractive index under photodynamic treatment was studied using OCT. Fat tissue slices 200-500 μm thick were used in in vitro experiments. To stain the fat tissue we used water-ethanol solutions of indocyanine green (ICG) and brilliant green (BG) with the concentration 1 mg/ml and 6 mg/ml, respectively. The CW laser diode (VD-VII DPSS, 808 nm) and the dental diode irradiator Ultra Lume Led 5 (442 and 597 nm) were used for irradiation of tissue slices. The irradiation time was 5 min for the laser and 15 min for the diode lamp. The experiments were carried out at room temperature. It was discovered the immersion optical clearing of fat tissue slices due to fat cell lipolysis under photodynamic treatment. Released cell content works as an immersion agent, thus the relative refractive index of tissue scatterers decreasing with the time elapsed after the treatment. These data support the hypothesis that photodynamic treatment induces fat cell lipolysis for some period after treatment.