This PDF file contains the editorial “Review Speed” for JBO Vol. 15 Issue 03

The significance of beam condition for scattered light from random tissue is analyzed for a practical optical imaging system with a finite numerical aperture. It is shown that in the transmitted illumination case, the information carrying part of the spectrum of the scattered light is proportional to the square of the spatial frequency of the spatial correlation function of the scattering potential of the medium. The result may be helpful in interpreting images obtained with microscopes in biological studies.

We introduce a new interferometric setup for single-exposure wide-field holographic phase imaging of highly dynamic biological samples. In this setup, the interferometric signal originates from a specially designed reflective interferometric chamber (InCh), creating an off-axis interferogram on the output plane of the system. The setup only requires the InCh and a simple reflection-mode two lens imaging system, without the need for additional optical elements such as gratings in the beam path. In addition, due to the close-to-common-path geometry of the setup, phase noise is greatly reduced. We experimentally compare the inherent phase stability of the system in ambient conditions to that of a conventional interferometer. We also demonstrate use of this system for wide-field quantitative phase imaging of two different highly dynamic, optically transparent biological samples: beating myocardial cells and moving unicellular microorganisms.

This work reports a miniaturized laparoscopic zoom camera that can significantly improve vision for minimally invasive surgery (MIS), also known as laparoscopic surgery. The laparoscopic zoom camera contains bioinspired fluidic lenses that can change curvature and focal length in a manner similar to the crystalline lenses in human eyes. The traditional laparoscope is long, rigid, and made of fixed glass lenses with a fixed field of view. The constricted vision of a laparoscope is often an inconvenience and plays a role in many surgical injuries. To further advance MIS technology, we developed a new type of laparoscopic camera that has a total length of less than 17 mm, greater than 4× optical zoom, and 100 times higher sensitivity than today's laparoscope allowing it to work under illumination as low as 300 lux. All these unique features are enabled by the technology of bioinspired fluidic lenses having a dynamic range over 100 diopters and being convertible between a convex and concave shape.

While multiphoton microscopy (MPM) has been performed with a wide range of excitation wavelengths, fluorescence emission has been limited to the visible spectrum. We introduce a paradigm for MPM of near-infrared (NIR) fluorescent molecular probes via nonlinear excitation at 1550 nm. This all-NIR system expands the range of available MPM fluorophores, virtually eliminates background autofluorescence, and allows for use of fiber-based, turnkey ultrafast lasers developed for telecommunications.

This work presents a novel tissue-mimicking phantom for use in a range of optical coherence tomography (OCT) experiments. Such phantoms are critical in the development and assessment of new OCT techniques, but no previously published phantoms have become universally accepted. We present the first description of a phantom based on a fibrin matrix, which improves key attributes of previously published methods. It provides a biocompatible, optically transparent scaffold in which to incorporate organic and/or inorganic optical scattering materials. Its fabrication time is markedly shorter than many common phantoms, and its lifetime is longer than other biocompatible phantoms. The potential of fibrin phantoms incorporating Intralipid^TM to introduce uniform optical scattering is demonstrated. The measured attenuation coefficient as a function of Intralipid concentration confirms the ability to control optical scattering. A bilayer phantom with distinct optical scattering in each layer is also presented.

We demonstrate for the first time that the detailed blood flow distribution within intracranial dura mater and cortex can be visualized by an ultrahigh sensitive optical microangiography (UHS-OMAG). The study uses an UHS-OMAG system operating at 1310 nm with an imaging speed at 150 frames per second that requires ~10 s to complete one 3-D scan of ~2.5×2.5 mm². The system is sensitive to blood flow with a velocity ranging from ~4 µm/s to ~23 mm/s. We show superior performance of UHS-OMAG in providing functional images of capillary level microcirculation within meninges in mice with the cranium left intact, the results of which correlate well with the standard dural histopathology.

Fourier domain optical coherence tomography (FD-OCT) requires either a linear-in-wavenumber spectrometer or a computationally heavy software algorithm to recalibrate the acquired optical signal from wavelength to wavenumber. The first method is sensitive to the position of the prism in the spectrometer, while the second method drastically slows down the system speed when it is implemented on a serially oriented central processing unit. We implement the full resampling process on a commercial graphics processing unit (GPU), distributing the necessary calculations to many stream processors that operate in parallel. A comparison between several recalibration methods is made in terms of performance and image quality. The GPU is also used to accelerate the fast Fourier transform (FFT) and to remove the background noise, thereby achieving full GPU-based signal processing without the need for extra resampling hardware. A display rate of 25 frames/sec is achieved for processed images (1024×1024 pixels) using a line-scan charge-coupled device (CCD) camera operating at 25.6 kHz.

We use laser damage thresholds in an in-vitro retinal model, and computational simulations to examine the laser exposure durations at which damage transitions from photothermal to photochemical at 413 nm. Our results indicate a dramatic shift in 1-h damage thresholds between exposure durations of 60 and 100 s. The trend in our in-vitro results is similar to a trend found in a recent study where retinal lesions were assessed 1-h post laser exposure in the rhesus eye Our data suggest that nonthermal mechanisms did not significantly contribute to cell death, even for exposures of 60 s. Knowledge of the transition point, and lack of concurrent thermal and nonthermal damage processes, are significant for those wishing to devise a comprehensive computational damage model.

An approach to quantitatively image targeted-agent binding rate in vivo is demonstrated with dual-probe injection of both targeted and nontargeted fluorescent dyes. Images of a binding rate constant are created that reveal lower than expected uptake of epidermal growth factor in an orthotopic xenograft pancreas tumor (2.3×10−5 s−1), as compared to the normal pancreas (3.4×10−5 s−1). This approach allows noninvasive assessment of tumor receptor targeting in vivo to determine the expected contrast, spatial localization, and efficacy in therapeutic agent delivery.

Dynamic speckle from 3-D coherence-gated optical sections provides a sensitive label-free measure of cellular activity up to 1 mm deep in living tissue. However, specificity to cellular functionality has not previously been demonstrated. In this work, we perform fluctuation spectroscopy on dynamic light scattering captured using coherence-domain digital holography to obtain the spectral response of tissue that is perturbed by temperature, osmolarity, and antimitotic cytoskeletal drugs. Different perturbations induce specific spectrogram response signatures that can show simultaneous enhancement and suppression in different spectral ranges.

We synthesize multifunctional microbubbles (MBs) for targeted delivery of antivascular endothelial growth factor (antiVEGF) therapy with multimodal imaging guidance. Poly-lactic-co-glycolic acid (PLGA) MBs encapsulating Texas Red dye are fabricated by a modified double-emulsion process. Simultaneous ultrasound and fluorescence imaging are achieved using Texas Red encapsulated MBs. The MBs are conjugated with Avastin, an antiVEGF antibody for treating neovascular age-related macular degeneration (AMD). The conjugation efficiency is characterized by enzyme-linked immunosorbent assay (ELISA). The efficiency for targeted binding of Avastin-conjugated MBs is characterized by microscopic imaging. Our work demonstrates the technical potential of using multifunctional MBs for targeted delivery of antiVEGF therapy in the treatment of exudative AMD.

Radio-frequency ablation (rfa) is the standard of care for the treatment of cardiac arrhythmias; however, there are no direct measures of the successful delivery of ablation lesions. Optical coherence tomography (OCT) imaging has the potential to provide real-time monitoring of cardiac rfa therapy, visualizing lesion formation and assessing tissue contact in the presence of blood. A rfa-compatible forward-imaging conical scanning probe is prototyped to meet this need. The forward-imaging probe provides circular scanning, with a 2-mm scan diameter and 30-µm spot size. During the application of rf energy, dynamics are recorded at 20 frames per second with a 40-kHz A-line rate. Real-time monitoring of cardiac rfa lesion formation and imaging in the presence of blood is demonstrated ex vivo in a swine left ventricle with a forward, flexible, circular scanning OCT catheter.

We present the application of a microfluidic optical cell stretcher to measure the elasticity of malaria-infected red blood cells. The measurements confirm an increase in host cell rigidity during the maturation of the parasite Plasmodium falciparum. The device combines the selectivity and sensitivity of single-cell elasticity measurements with a throughput that is higher than conventional single-cell techniques. The method has potential to detect early stages of infection with excellent sensitivity and high speed.

Astrocytes have been identified to actively contribute to brain functions through Ca²⁺ signaling, serving as a bridge to communicate with neurons and other brain cells. However, conventional stimulation techniques are hard to apply to delicate investigations on astrocytes. Our group previously reported photostimulation with a femtosecond laser to evoke astrocytic calcium (Ca²⁺) waves, providing a noninvasive and efficient approach with highly precise targeting. In this work, detailed characteristics of astrocytic Ca²⁺ signaling induced by photostimulation are presented. In a purified astrocytic culture, after the illumination of a femtosecond laser onto one cell, a Ca²⁺ wave throughout the network with reduced speed is induced, and intracellular Ca²⁺ oscillations are observed. The intercellular propagation is pharmacologically confirmed to be mainly mediated by ATP through P2Y receptors. Different patterns of Ca²⁺ elevations with increased amplitude in the stimulated astrocyte are discovered by varying the femtosecond laser power, which is correspondingly followed by broader intercellular waves. These indicate that the strength of photogenerated Ca²⁺ signaling in astrocytes has a positive relationship with the stimulating laser power. Therefore, distinct Ca²⁺ signaling is feasibly available for specific studies on astrocytes by employing precisely controlled photostimulation.

The study of light propagation in diffusive media requires solving the radiative transfer equation, or eventually, the diffusion approximation. Except for some cases involving simple geometries, the problem with immersed inclusions has not been solved. Also, Monte Carlo (MC) calculations have become a gold standard for simulating photon migration in turbid media, although they have the drawback large processing times. The purpose of this work is two-fold: first, we introduce a new processing criterion to retrieve information about the location and shape of absorbing inclusions based on normalization to the background intensity, when no inhomogeneities are present. Second, we demonstrate the feasibility of including inhomogeneities in MC simulations implemented in graphics processing units, achieving large acceleration factors (~10³), thus providing an important tool for iteratively solving the forward problem to retrieve the optical properties of the inclusion. Results using a cw source are compared with MC outcomes showing very good agreement.

Transillumination (TI) of dental enamel with near-infrared light is a promising nonionizing imaging method for detection of early caries lesion. Increased mineral loss (caries lesion) leads to increased scattering and absorption. Caries thus appear as dark regions because less light reaches the detector. The aim of this work was to characterize the performance of a TI system from the resolution of acquired images using the modulation transfer function at two wavelengths, 1.28 and 1.4 µm. Test charts with various values of spatial periods, mimicking a perfect caries lesion, were attached to tooth sections, followed by capture of the transmitted image, using both wavelengths. The sections were then consecutively reduced in thickness, and a sequence of all sizes of the test charts were used for repeatedly imaging procedures. The results show that the TI system can detect feature size of 250 µm with 30% modulation. From the information about how the image degrades as it propagates through enamel, we also examined the possibility of estimating the position of a simulated approximal caries lesion by comparing images obtained from the two sides of a tooth section.

Conventionally, liver fibrosis is diagnosed using histopathological techniques. The traditional method is time-consuming in that the specimen preparation procedure requires sample fixation, slicing, and labeling. Our goal is to apply multiphoton microscopy to efficiently image and quantitatively analyze liver fibrosis specimens bypassing steps required in histological preparation. In this work, the combined imaging modality of multiphoton autofluorescence (MAF) and second-harmonic generation (SHG) was used for the qualitative imaging of liver fibrosis of different METAVIR grades under label-free, ex vivo conditions. We found that while MAF is effective in identifying cellular architecture in the liver specimens, it is the spectrally distinct SHG signal that allows the characterization of the extent of fibrosis. We found that qualitative SHG imaging can be used for the effective identification of the associated features of liver fibrosis specimens graded METAVIR 0 to 4. In addition, we attempted to associate quantitative SHG signal to the different METAVIR grades and found that an objective determination of the extent of disease progression can be made. Our approach demonstrates the potential of using multiphoton imaging in rapid classification of ex vivo liver fibrosis in the clinical setting and investigation of liver fibrosis-associated physiopathology in animal models in vivo.

When a large, highly absorbing breast lesion is imaged by optical tomography in reflection geometry, most of the photons are absorbed by the top portion of the lesion. As a result, the lower portion of the lesion is not quantified correctly. This posterior light shadowing effect is similar to the sound shadowing effect frequently seen in pulse-echo ultrasound images. The presence of significant posterior shadowing of a lesion in ultrasound images suggests malignance. The light shadowing effect due to optical contrast is characterized using a simple measure and validated by the Monte Carlo photon-tracking method and phantom experiments. Clinical examples of large malignant and benign lesions are presented to demonstrate the shadowing effect and the utility of the measure. Understanding and quantifying the shadowing effect due to optical contrast is important for characterizing larger malignant cancers from benign lesions.

Solid tumors are characterized by abnormal blood vessel organization, structure, and function. These abnormalities give rise to enhanced vascular permeability and may predict therapeutic responses. The permeability and architecture of the microvasculature in human osteosarcoma tumors growing in dorsal window chambers in athymic mice were measured by confocal laser scanning microscopy (CLSM) and dynamic contrast enhanced magnetic resonance imaging (DCE-MRI). Dextran (40 kDa) and Gadomer were used as molecular tracers for CLSM and DCE-MRI, respectively. A significant correlation was found between permeability indicators. The extravasation rate K_i as measured by CLSM correlated positively with DCE-MRI parameters, such as the volume transfer constant K^trans and the initial slope of the contrast agent concentration-time curve. This demonstrates that these two techniques give complementary information. Extravasation was further related to microvascular structure and was found to correlate with the fractal dimension and vascular density. The structural parameter values that were obtained from CLSM images were higher for abnormal tumor vasculature than for normal vessels.

We applied multiphoton autofluorescence (MAF) and second-harmonic generation (SHG) microscopy to monitor corneal wound healing after photorefractive keratectomy (PRK). Our results show that keratocyte activation can be observed by an increase in its MAF, while SHG imaging of corneal stroma can show the depletion of Bowman's layer after PRK and the reticular collagen deposition in the wound healing stage. Furthermore, quantification of the keratocyte activation and collagen deposition in conjunction with immunohistochemistry and histological images demonstrate the effectiveness of mitomycin C (MMC) in suppressing myofibroblast proliferation and collagen regeneration in the post-PRK wound healing process.

The recent development of hybrid imaging scanners that integrate fluorescence molecular tomography (FMT) and x-ray computed tomography (XCT) allows the utilization of x-ray information as image priors for improving optical tomography reconstruction. To fully capitalize on this capacity, we consider a framework for the automatic and fast detection of different anatomic structures in murine XCT images. To accurately differentiate between different structures such as bone, lung, and heart, a combination of image processing steps including thresholding, seed growing, and signal detection are found to offer optimal segmentation performance. The algorithm and its utilization in an inverse FMT scheme that uses priors is demonstrated on mouse images.

Affibody protein is an engineered protein scaffold with a three-helical bundle structure. Affibody molecules of small size (7 kD) have great potential for targeting overexpressed cancer biomarkers in vivo. To develop an Affibody-based molecular probe for in vivo optical imaging of epidermal growth factor receptor (EGFR) positive tumors, an anti-EGFR Affibody molecule, Ac-Cys-Z_EGFR:1907 (7 kD), is site-specifically conjugated with a near-IR fluorescence dye, Cy5.5-mono-maleimide. Using fluorescent microscopy, the binding specificity of the probe Cy5.5-Z_EGFR:1907 is checked by a high-EGFR-expressing A431 cell and low-EGFR-expressing MCF7 cells. The binding affinity of Cy5.5-Z_EGFR:1907 (K_D) to EGFR is 43.6±8.4 nM, as determined by flow cytometry. For an in vivo imaging study, the probe shows fast tumor targeting and good tumor contrast as early as 0.5 h postinjection (p.i.) for A431 tumors, while MCF7 tumors are barely visible. An ex vivo imaging study also demonstrates that Cy5.5-Z_EGFR:1907 has high tumor, liver, and kidney uptakes at 24 h p.i.. In conclusion, Cy5.5-Z_EGFR:1907 shows good affinity and high specificity to the EGFR. There is rapid achievement of good tumor-to-normal-tissue contrasts of Cy5.5-Z_EGFR:1907, thus demonstrating its potential for EGFR-targeted molecular imaging of cancers.

We demonstrate the utility of functional near-infrared spectroscopy (fNIRS) as a tool for physicians to study cortical plasticity in children with cerebral palsy (CP). Motor cortex activation patterns were studied in five healthy children and five children with CP (8.4±2.3 years old in both groups) performing a finger-tapping protocol. Spatial (distance from center and area difference) and temporal (duration and time-to-peak) image metrics are proposed as potential biomarkers for differentiating abnormal cortical activation in children with CP from healthy pediatric controls. In addition, a similarity image-analysis concept is presented that unveils areas that have similar activation patterns as that of the maximum activation area, but are not discernible by visual inspection of standard activation images. Metrics derived from the images presenting areas of similarity are shown to be sensitive identifiers of abnormal activation patterns in children with CP. Importantly, the proposed similarity concept and related metrics may be applicable to other studies for the identification of cortical activation patterns by fNIRS.

Digital holographic microscopy (DHM) enables quantitative multifocus phase contrast imaging for nondestructive technical inspection and live cell analysis. Time-lapse investigations on human brain microvascular endothelial cells demonstrate the use of DHM for label-free dynamic quantitative monitoring of cell division of mother cells into daughter cells. Cytokinetic DHM analysis provides future applications in toxicology and cancer research.

Predicting tumor metastatic potential remains a challenge in cancer research and clinical practice. Our goal was to identify novel biomarkers for differentiating human breast tumors with different metastatic potentials by imaging the in vivo mitochondrial redox states of tumor tissues. The more metastatic (aggressive) MDA-MB-231 and less metastatic (indolent) MCF-7 human breast cancer mouse xenografts were imaged with the low-temperature redox scanner to obtain multi-slice fluorescence images of reduced nicotinamide adenine dinucleotide (NADH) and oxidized flavoproteins (Fp). The nominal concentrations of NADH and Fp in tissue were measured using reference standards and used to calculate the Fp redox ratio, Fp/(NADH+Fp). We observed significant core-rim differences, with the core being more oxidized than the rim in all aggressive tumors but not in the indolent tumors. These results are consistent with our previous observations on human melanoma mouse xenografts, indicating that mitochondrial redox imaging potentially provides sensitive markers for distinguishing aggressive from indolent breast tumor xenografts. Mitochondrial redox imaging can be clinically implemented utilizing cryogenic biopsy specimens and is useful for drug development and for clinical diagnosis of breast cancer.

Diagnosing sinusitis remains a challenge for primary care physicians. There is a need for a simple, office-based technique to aid in the diagnosis of sinusitis without the cost and radiation risk of conventional radiologic imaging. We designed a low-cost near-infrared (NIR) device to transilluminate the maxillary sinuses. The use of NIR light allows for greater interrogation of deep-tissue structures as compared to visible light. NIR imaging of 21 patients was performed and compared with computed tomography (CT) scans. Individual maxillary sinuses were scored on a scale from 0 to 2 based on their degree of aeration present on CT and similarly based on the NIR signal penetration into the maxilla on NIR images. Our results showed that air-filled and fluid/tissue-filled spaces can be reasonably distinguished by their differing NIR signal penetration patterns, with average NIR imaging scores for fluid-filled maxillary sinuses (0.93±0.78, n=29) significantly lower than those for normal maxillary sinuses (1.62±0.57, n=13) (p=0.003). NIR imaging of the sinuses is a simple, safe, and cost-effective modality that can potentially aid in the diagnosis of sinusitis. Long-term, significant device refinement and large clinical trials will be needed to determine the diagnostic accuracy of this technique.

Previous studies have demonstrated the ultrasound-induced skin optical clearing enhancement with topical application of 60% glycerol (G) on in vitro porcine skin and in vivo human skin. Our purpose was to find the relation between the effect of optical skin clearing and different concentrations of glycerol and to find more effective ultrasound-glycerol combinations on optical skin clearing. The enhancement effect of ultrasound [Sonophoresis (SP) delivery] in combination with 40% G, 60% G, and 80% G on in vitro human skin optical clearing was investigated. Light imaging depths of skin were measured using optical coherence tomography. Different concentrations of glycerol and ultrasound with a frequency of 1 MHz and an intensity of 0.5 W/cm2 was simultaneously applied for 15 min. The results show that with the increase of concentration of glycerol, the optical clearing of skin is much improved. Optical clearing capability of glycerol was more enhanced with simultaneous application of ultrasound compared with glycerol alone. The attenuation coefficients of skin tissues after application of 40% G/SP, 60% G/SP, and 80% G/SP decreased approximately 11.8%, 18.5%, and 20.0% at 15 min compared with 40% G, 60% G, and 80% G alone, respectively. The greatest decrease in attenuation coefficients at 60 min was approximately 52.3% and 63.4% for 80% G (without ultrasound) and 80% G/SP (with ultrasound), respectively, which are 2.1-fold and 2.6-fold to that in the 40% G.

Sinusoidally structured illumination is used in concert with a phantom-based lookup-table (LUT) to map wide-field optical properties in turbid media with reduced albedos as low as 0.44. A key advantage of the lookup-table approach is the ability to measure the absorption (µa) and reduced scattering coefficients (µ′_s) over a much broader range of values than permitted by current diffusion theory methods. Through calibration with a single reflectance standard, the LUT can extract µ′_s from 0.8 to 2.4 mm^-1 with an average root-mean-square (rms) error of 7% and extract µ_a from 0 to 1.0 mm^-1 with an average rms error of 6%. The LUT is based solely on measurements of two parameters, reflectance R and modulation M at an illumination period of 10 mm. A single set of three phase-shifted images is sufficient to measure both M and R, which are then used to generate maps of absorption and scattering by referencing the LUT. We establish empirically that each pair (M,R) maps uniquely to only one pair of (µ′_s,µ_a) and report that the phase function (i.e., size) of the scatterers can influence the accuracy of optical property extraction.

A novel rotating linear polarization imaging technique is developed to characterize the anisotropic properties of tissues. Differences of orthogonal linear polarization with different incident and detection polarization angles are fitted to an analytical function to retrieve a set of parameters. Experiments with different tissues and Monte Carlo simulations indicate that two of the parameters, G and φ₃/2, are correlated to the anisotropic property and the orientation angle of the fibrous structure in the media. The technique can be used for clinical diagnosis.

Gold nanoparticles (AuNPs) have the potential to become a versatile biomarker. For further use of AuNPs labeled with functionalized molecules, their visualization in biological systems by routine laboratory tools such as light microscopy is crucial. However, the size far below the diffraction limit affords specialized parameters for microscopical detection, which stimulated the current study, aimed to determine from which size onward AuNPs, either in dispersion or cell-associated, can be reliably detected by standard confocal microscopy. First, gold colloids of size-restricted fractions are examined in dispersion. At a minimum particle size of 60 nm, detection appears to be reliable. Particle counts in dilution series confirm these results by revealing single particle detection of 60-nm colloids. Second, AuNPs are visualized and quantified in cells, which interestingly cause a phase shift in the reflection of AuNPs. Gold mass spectroscopy confirms the number of AuNPs counted microscopically inside cells. Furthermore, it demonstrates for the first time a very high diffusion rate of 15-nm particles into the cells. In conclusion, the results back the suitability of confocal microscopy for the quantitative tracking of colloidal and intracellular gold nanoparticles sized 60 nm.

In this feasibility study, we present a method for virtual 4-D imaging of healthy and injured subpleural lung tissue in the ventilated mouse. We use triggered swept source optical coherence tomography (OCT) with an A-scan frequency of 20 kHz to image murine subpleural alveoli during the inspiratory phase. The data acquisition is gated to the ventilation pressure to take single B-scans in each respiration cycle for different pressure levels. The acquired B-scans are combined off-line into one volume scan for each pressure level. The air fraction in healthy lungs and injured lungs is measured using 2-D OCT en-face images. Upon lung inspiration from 2 to 12 cmH₂O ventilation pressure, the air fraction increases in healthy lungs by up to 11% and in injured lungs by 8%. This expansion correlates well with results of previous studies, reporting increased alveolar area with increased ventilation pressures. We demonstrate that OCT is a useful tool to investigate alveolar dynamics in spatial dimensions.

Typical imaging depths with multiphoton microscopy (MPM) are limited to less than 300 µm in many tissues due to light scattering. Optical clearing significantly reduces light scattering by replacing water in the organ tissue with a fluid having a similar index of refraction to that of proteins. We demonstrate MPM of intact, fixed, cleared mouse organs with penetration depths and fields of view in excess of 2 mm. MPM enables the creation of large 3-D data sets with flexibility in pixel format and ready access to intrinsic fluorescence and second-harmonic generation. We present high-resolution images and 3-D image stacks of the brain, small intestine, large intestine, kidney, lung, and testicle with image sizes as large as 4096×4096 pixels.

Planar glass waveguides with a specific number of modes were fabricated by Ag⁺-Na⁺ exchange in Schott SG11 glass. The effective refractive indices were determined using m-line spectroscopy in both s- and p-polarization. By using the reversed Wentzel-Kramers-Brillouin approximation, the index profiles were described by a nonlinear diffusion equation. The diffusion coefficients for Ag⁺ were established, as well as the penetration depth of the evanescent field in an aqueous environment for the different modes. The integrals of |E|² fields for the evanescent-guided fields were investigated. These are important when evanescent fields are used for illumination in interface microscopy, an alternative method to total internal reflection fluorescence (TIRF) microscopy. The photoluminescent behavior of the waveguides was investigated as a function of ion exchange time and excitation wavelengths. Comparable images were obtained of fluorescently labeled HEK293 cells using TIRF microscopy and waveguide evanescent field fluorescence microscopy. Imaging was performed using HEK293 cells, delivering similar images and information.

Photoacoustic tomography (PAT) has been widely used to image optically absorptive objects in both human and animal tissues. For the first time, we present imaging of water with laser-based PAT. We photoacoustically measure the absorption spectra of water-ethanol mixtures at various water concentrations, and then image water-ethanol and pure-water inclusions in gel and a water inclusion in fat tissue. The significant difference in photoacoustic signals between water and fat tissue indicates that the laser-based PAT has the potential to detect water content in tissue.

We develop a fiber optical coherence tomography (OCT) system in the clinical utility of imaging port wine stains (PWS). We use our OCT system on 41 patients with PWS to document the difference between PWS skin and contralateral normal skin. The system, which operates at 4 frames/s with axial and transverse resolutions of 10 and 9 µm, respectively, in the skin tissue, can clearly distinguish the dilated dermal blood vessels from normal tissue. We present OCT images of patients with PWS and normal human skin. We obtain the structural parameters, including epidermal thickness and diameter and depth of dilated blood vessels. We demonstrate that OCT may be a useful tool for the noninvasive imaging of PWS. It may help determine the photosensitizer dose and laser parameters in photodynamic therapy for treating port wine stains.

Integrated Raman and angular-scattering microscopy (IRAM) is a multimodal platform capable of noninvasively probing both the chemistry and morphology of a single cell without prior labeling. Using this system, we are able to detect activation-dependent changes in the Raman and elastic-scattering signals from CD8+ T cells stimulated with either Staphylococcal enterotoxin B (SEB) or phorbol myristate acetate (PMA). In both cases, results obtained from the IRAM instrument correlate well with results obtained from traditional fluorescence-based flow cytometry for paired samples. SEB-mediated activation was distinguished from resting state in CD8+ T cells by an increase in the number and mean size of small (~500-nm) elastic scatterers as well as a decrease in Raman bands, indicating changes in nuclear content. PMA-mediated activation induced a different profile in CD8+ T cells from SEB, showing a similar increase in small elastic scatterers but a different Raman change, with elevation of cellular protein and lipid bands. These results suggest the potential of this multimodal, label-free optical technique for studying processes in single cells.

We present the theoretical background and experimental validation of a pupil tracking method based on measurement of the irradiance centroid of Hartmann-Shack aberrometric images. The experimental setup consists of a Hartmann-Shack (HS) sensor forming over the same camera the images of the eye's pupil and the aberrometric image. The calibration is made by comparing the controlled displacements induced to an artificial eye with the displacements estimated from the centroid of the pupil and of the HS focal plane. The pupil image is also used for validation of the method when operating with human eyes. The experimental results after calibration show a root mean square error of 10.45 µm for the artificial eye and 27, 10, and 6 µm rms for human eyes tested using Hartmann-Shack images, with signal-to-noise ratios of 6, 8, and 11, respectively. The performance of the method is similar to conventional commercial eye trackers. It avoids the need for using separate tracking devices and their associated synchronization problems. This technique can also be used to reprocess present and stored sets of Hartmann-Shack aberrometric images to estimate the ocular movements that occurred during the measurement runs, and, if convenient, to correct the measured aberrations from their influence.

There is a demand for technologies able to assess the perfusion of surgical flaps quantitatively and reliably to avoid ischemic complications. The aim of this study is to test a new high-speed high-definition laser Doppler imaging (LDI) system (FluxEXPLORER, Microvascular Imaging, Lausanne, Switzerland) in terms of preoperative mapping of the vascular supply (perforator vessels) and postoperative flow monitoring. The FluxEXPLORER performs perfusion mapping of an area 9×9 cm with a resolution of 256×256 pixels within 6 s in high-definition imaging mode. The sensitivity and predictability to localize perforators is expressed by the coincidence of preoperatively assessed LDI high flow spots with intraoperatively verified perforators in nine patients. 18 free flaps are monitored before, during, and after total ischemia. 63% of all verified perforators correspond to a high flow spot, and 38% of all high flow spots correspond to a verified perforator (positive predictive value). All perfused flaps reveal a value of above 221 perfusion units (PUs), and all values obtained in the ischemic flaps are beneath 187 PU. In summary, we conclude that the present LDI system can serve as a reliable, fast, and easy-to-handle tool to detect ischemia in free flaps, whereas perforator vessels cannot be detected appropriately.

There is considerable interest in developing new methods for in vivo imaging of the complex anatomy of the mammalian cochlea for clinical as well as fundamental studies. In this study, we explored, the feasibility of spectral domain optical coherence tomography (SD-OCT) for 3-D in vivo imaging of the cochlea in mice. The SD-OCT system employed in this study used a broadband light source centered at 1300 nm, and the imaging speed of the system was 47,000 A-scans per second using the InGaAs camera. The system was capable of providing fully processed, high-resolution B-scan images [512 (axial)×128 (lateral) pixels] at 280 frames per sec. The 3-D imaging acquisition time for a whole cochlea was ~0.45 sec. The traditional SD-OCT structural imaging algorithm was used to reconstruct 3-D cochlear morphology. We demonstrated that SD-OCT can be successfully used for in vivo imaging of important morphological features within the mouse cochlea, such as the otic capsule and structures within, including Reissner's membrane, the basilar membrane, tectorial membrane, organ of Corti, and modiolus of the apical and middle turns.

An automated method is presented for artery-vein separation in cerebral cortical images recorded with optical imaging of the intrinsic signal. The vessel-type separation method is based on the fact that the spectral distribution of intrinsic physiological oscillations varies from arterial regions to venous regions. In arterial regions, the spectral power is higher in the heartbeat frequency (HF), whereas in venous regions, the spectral power is higher in the respiration frequency (RF). The separation method was begun by extracting the vascular network and its centerline. Then the spectra of the optical intrinsic signals were estimated by the multitaper method. A standard F-test was performed on each discrete frequency point to test the statistical significance at the given level. Four periodic physiological oscillations were examined: HF, RF, and two other eigenfrequencies termed F1 and F2. The separation of arteries and veins was implemented with the fuzzy c-means clustering method and the region-growing approach by utilizing the spectral amplitudes and power-ratio values of the four eigenfrequencies on the vasculature. Subsequently, independent spectral distributions in the arteries, veins, and capillary bed were estimated for comparison, which showed that the spectral distributions of the intrinsic signals were very distinct between the arterial and venous regions.

This study investigates differences in the response of breast tumor tissue versus healthy fibroglandular tissue to inspired gases. Cycles of carbogen and oxygen gas are administered while measuring the changes with magnetic-resonance-guided near-infrared imaging in a pilot study of breast cancers. For two patients, analyses are performed with cross-correlation techniques, which measure the strength of hemodynamic modulation. The results show that the overall vasoresponse, indicated by total hemoglobin, of healthy tissue has approximately a 72% and 41% greater correlation to the gas stimulus than the tumor region, in two patients respectively, when background physiological changes are controlled. These data support the hypothesis that tumor vasculature has a poorly functioning vasodilatory mechanism, most likely caused by dysfunctional smooth muscle cells lining the vasculature. This study presents a methodology to quantitatively analyze inspired gas changes in human breast tumors, and demonstrates this technique in a pilot patient population.

Transgenic reporter mice and advances in imaging instrumentation are enabling real-time visualization of cellular mechanisms in living subjects and accelerating the development of novel therapies. Innovative confocal microscope designs are improving their utility for microscopic imaging of fluorescent reporters in living animals. We develop dual-axis confocal DAC microscopes for such in vivo studies and create mouse models where fluorescent proteins are expressed in the skin for the purpose of advancing skin therapeutics and transdermal delivery tools. Three-dimensional image volumes, through the different skin compartments of the epidermis and dermis, can be acquired in several seconds with the DAC microscope in living mice, and are comparable to histologic analyses of reporter protein expression patterns in skin sections. Intravital imaging with the DAC microscope further enables visualization of green fluorescent protein GFP reporter gene expression in the skin over time, and quantification of transdermal delivery of small interfering RNA siRNA and therapeutic efficacy. Visualization of transdermal de-livery of nucleic acids will play an important role in the development of innovative strategies for treating skin pathologies.

Polarized Raman spectroscopy allows measurement of molecular orientation and composition and is widely used in the study of polymer systems. Here, we extend the technique to the extraction of quantitative orientation information from bone tissue, which is optically thick and highly turbid. We discuss multiple scattering effects in tissue and show that repeated measurements using a series of objectives of differing numerical apertures can be employed to assess the contributions of sample turbidity and depth of field on polarized Raman measurements. A high numerical aperture objective minimizes the systematic errors introduced by multiple scattering. We test and validate the use of polarized Raman spectroscopy using wild-type and genetically modified (oim/oim model of osteogenesis imperfecta) murine bones. Mineral orientation distribution functions show that mineral crystallites are not as well aligned (p<0.05) in oim/oim bones (28±3 deg) compared to wild-type bones (22±3 deg), in agreement with small-angle X-ray scattering results. In wild-type mice, backbone carbonyl orientation is 76±2 deg and in oim/oim mice, it is 72±4 deg (p>0.05). We provide evidence that simultaneous quantitative measurements of mineral and collagen orientations on intact bone specimens are possible using polarized Raman spectroscopy.

Coherence between spontaneous fluctuations in arterial blood pressure (ABP) and the cerebral near-infrared spectroscopy signal can detect cerebral autoregulation. Because reliable measurement depends on signals with high signal-to-noise ratio, we hypothesized that coherence is more precisely determined when fluctuations in ABP are large rather than small. Therefore, we investigated whether adjusting for variability in ABP (variability_ABP) improves precision. We examined the impact of variability_ABP within the power spectrum in each measurement and between repeated measurements in preterm infants. We also examined total monitoring time required to discriminate among infants with a simulation study. We studied 22 preterm infants (GA<30) yielding 215 10-min measurements. Surprisingly, adjusting for variability_ABP within the power spectrum did not improve the precision. However, adjusting for the variability_ABP among repeated measurements (i.e., weighting measurements with high variability_ABP in favor of those with low) improved the precision. The evidence of drift in individual infants was weak. Minimum monitoring time needed to discriminate among infants was 1.3-3.7 h. Coherence analysis in low frequencies (0.04-0.1 Hz) had higher precision and statistically more power than in very low frequencies (0.003-0.04 Hz). In conclusion, a reliable detection of cerebral autoregulation takes hours and the precision is improved by adjusting for variability_ABP between repeated measurements.

We study the properties of single red blood cells (RBCs) held in an optical-tweezers trap. We observe a change in the spectrum of Brownian fluctuations between RBCs from normal and malaria-infected samples. The change, caused by infection-induced structural changes in the cell, appears as a statistical increase in the mean (by 25%) and standard deviation (by 200%) of the corner frequency measured over ~100 cells. The increase is observed even though the ensemble of cells being measured consists mostly of cells that do not actually host the parasite, but are from an infected pool. This bystander effect appears to vindicate other observations that infected cells can affect the biomechanical properties of uninfected cells. The change is also observed to be independent of the stage of infection and its duration, highlighting its potential for disease detection.

We employ a hybrid diffuse correlation spectroscopy (DCS) and near-infrared spectroscopy (NIRS) monitor for neonates with congenital heart disease (n=33). The NIRS-DCS device measured changes during hypercapnia of oxyhemoglobin, deoxyhemoglobin, and total hemoglobin concentrations; cerebral blood flow (rCBF_DCS); and oxygen metabolism (rCMRO₂). Concurrent measurements with arterial spin-labeled magnetic resonance imaging (rCBF_ASL-MRI, n=12) cross-validate rCBF_DCS against rCBF_ASL-MRI, showing good agreement (R=0.7, p=0.01). The study demonstrates use of NIRS-DCS on a critically ill neonatal population, and the results indicate that the optical technology is a promising clinical method for monitoring this population.

There are several noninvasive techniques for assessing the kinetics of tear film, but no comparative studies have been conducted to evaluate their efficacies. Our aim is to test and compare techniques based on high-speed videokeratoscopy (HSV), dynamic wavefront sensing (DWS), and lateral shearing interferometry (LSI). Algorithms are developed to estimate the tear film build-up time T_BLD, and the average tear film surface quality in the stable phase of the interblink interval TFSQ_Av. Moderate but significant correlations are found between T_BLD measured with LSI and DWS based on vertical coma (Pearson's r²=0.34, p<0.01) and higher order rms (r²=0.31, p<0.01), as well as between TFSQ_Av measured with LSI and HSV (r²=0.35, p<0.01), and between LSI and DWS based on the rms fit error (r²=0.40, p<0.01). No significant correlation is found between HSV and DWS. All three techniques estimate tear film build-up time to be below 2.5 sec, and they achieve a remarkably close median value of 0.7 sec. HSV appears to be the most precise method for measuring tear film surface quality. LSI appears to be the most sensitive method for analyzing tear film build-up.

We use near-infrared spectroscopy (NIRS) to assess listeners' cortical responses to a 10-s series of pure tones separated in frequency. Listeners are instructed to either judge the rhythm of these "streaming" stimuli (active-response listening) or to listen to the stimuli passively. Experiment 1 shows that active-response listening causes increases in oxygenated hemoglobin (oxy-Hb) in response to all stimuli, generally over the (pre)motor cortices. The oxy-Hb increases are significantly larger over the right hemisphere than over the left for the final 5 s of the stimulus. Hemodynamic levels do not vary with changes in the frequency separation between the tones and corresponding changes in perceived rhythm ("gallop," "streaming," or "ambiguous"). Experiment 2 shows that hemodynamic levels are strongly influenced by listening mode. For the majority of time windows, active-response listening causes significantly larger oxy-Hb increases than passive listening, significantly over the left hemisphere during the stimulus and over both hemispheres after the stimulus. This difference cannot be attributed to physical motor activity and preparation related to button pressing after stimulus end, because this is required in both listening modes.

We probe volar-side fingertip capillary beds with near-infrared laser light and collect Raman, Rayleigh, and Mie scattered light and fluorescence. The results are interpreted using radiation transfer theory in the single-scattering approximation. The surface topography of the skin is modeled using the Fresnel equations. The skin is treated as a three-layer material, with a mean-field treatment of tissue composition and related optical properties. The model, with a reasonable choice of tissue parameters, gives a remarkably accurate account of the features of actual measurements. It predicts the optimal values for the incident angle of the laser beam and the distance between beam and detector. It explains the correlated temporal changes in the intensities of elastically and inelastically scattered light caused by heart-driven pulses and why they are out of phase. With appropriate boundary conditions, the model can be used to discuss the scattering from ridged skin extruded conformally into an aperture in a metal surface under constant light pressure. The probing results suggest an inherent regularity and similarity in the anatomy and composition of surface and subsurface tissues of a wide range of skin types.

Identification of hematopoietic stem cells (HSCs) in different stages of maturation is one of the major issues in stem cell research and bone marrow (BM) transplantation. Each stage of maturation of HSCs is characterized by a series of distinct glycoproteins present on the cell plasma membrane surface, named a cluster of differentiation (CD). Currently, complicated and expensive procedures based on CD expression are needed for identification and isolation of HSCs. This method is under dispute, since the correct markers' composition is not strictly clear, thus there is need for a better method for stem cell characterization. In the present study, Fourier transform infrared (FTIR) spectroscopy is employed as a novel optical method for identification and characterization of HSCs based on their entire biochemical features. FTIR spectral analysis of isolated mice HSCs reveals several spectral markers related to lipids, nucleic acids, and carbohydrates, which distinguish HSCs from BM cells. The unique "open" conformation of HSC DNA as identified by FTIR is exploited for HSCs quantification in the BM. The proposed method of FTIR spectroscopy for HSC identification and quantification can contribute to stem cell research and BM transplantation.

We explore the feasibility of using combined fluorescence and diffuse reflectance spectroscopy to characterize a myocardial infarct at different developing stages. An animal study is conducted using rats with surgically induced myocaridal infarction (MI). In vivo fluorescence spectra at 337-nm excitation and diffuse reflectance between 400 and 900 nm are measured from the heart. Spectral acquisition is performed: 1. for normal heart tissue; 2. for the area immediately surrounding the infarct; and 3. for the infarcted tissue itself, one, two, three, and four weeks into MI development. Histological and statistical analyses are used to identify unique pathohistological features and spectral alterations associated with the investigated regions. The main alterations (p<0.05) in diffuse reflectance spectra are identified primarily between 450 and 600 nm. The dominant fluorescence alterations are increases in peak fluorescence intensity at 400 and 460 nm. The extent of these spectral alterations is related to the duration of the infarction. The findings of this study support the concept that optical spectroscopy could be useful as a tool to noninvasively determine the in vivo pathophysiological features of a myocardial infarct and its surrounding tissue, thereby providing real-time feedback to surgeons during various surgical interventions for MI.

We applied near-infrared spectroscopy (NIRS) and electroencephalography (EEG) simultaneously on the mouse brain and investigated the hemodynamic response to epileptic episodes under pharmacologically driven seizure. γ-butyrolactone (GBL) and 4-aminopyridine (4-AP) were applied to induce absence and tonic-clonic seizures, respectively. The epileptic episodes were identified from the single-channel EEG, and the corresponding hemodynamic changes in different regions of the brain were characterized by multichannel frequency-domain NIRS. Our results are the following: (i) the oxyhemoglobin level increases in the case of GBL-treated mice but not 4-AP-treated mice compared to the predrug state; (ii) the dominant response to each absence seizure is a decrease in deoxyhemolobin; (iii) the phase shift between oxy- and deoxyhemoglobin reduces in GBL-treated mice but no 4-AP-treated mice; and (iv) the spatial correlation of hemodynamics increased significantly in 4-AP-treated mice but not in GBL-treated mice. Our results shows that spatiotemporal tracking of cerebral hemodynamics using NIRS can be successfully applied to the mouse brain in conjunction with electrophysiological recording, which will support the study of molecular, cellular, and network origin of neurovascular coupling in vivo.

We experimentally study potential mechanisms by which the enhancement factor in low-coherence enhanced backscattering (LEBS) can probe subtle variations in radial intensity distribution in weakly scattering media. We use enhanced backscattering of light by implementing either (1) low spatial coherence illumination or (2) multiple spatially independent detections using a microlens array under spatially coherent illumination. We show that the enhancement factor in these configurations is a measure of the integrated intensity within the localized coherence or detection area, which can exhibit strong dependence on small perturbations in scattering properties. To further evaluate the utility of the LEBS enhancement factor, we use a well-established animal model of cutaneous two-stage chemical carcinogenesis. In this pilot study, we demonstrate that the LEBS enhancement factor can be substantially altered at a stage of preneoplasia. Our animal result supports the idea that early carcinogenesis can cause subtle alterations in the scattering properties that can be captured by the LEBS enhancement factor. Thus, the LEBS enhancement factor has the potential as an easily measurable biomarker in skin carcinogenesis.

Optical polarimetry for monitoring glucose concentration in the aqueous humor of the eye as a potential noninvasive means of assessing blood glucose has promise, but the realization of such an approach has been limited by noise from time-varying corneal birefringence due to motion artifact. Modeling the corneal birefringence of the eye is critically important toward understanding the overall effect of this noise source compared to other changes in the signal, and can aid in design of the polarimetric system. To this end, an eye model is introduced in this work that includes spatially varying birefringence properties of the cornea. The degree of birefringence and the fast axis orientation is calculated as a function of beam position on the anterior chamber. It is shown that the minimum change in polarization vector orientation occurs for beam position near the midpoint between the corneal apex and limbus. In addition, the relative wavelength independence of motion artifact is shown in the same region. The direct consequence of these findings are that a multiwavelength polarimetric system can potentially be utilized to eliminate the effect of time-varying corneal birefringence, and that eye coupling is optimal at the midpoint between the apex and limbus.

Reduced nicotinamide adenine dinucleotide (NADH) is a well-known metabolic coenzyme and endogenous fluorophore. In this study, we develop a system that simultaneously measures time- and wavelength-resolved fluorescence to extract free and protein-bound NADH signals from total cellular fluorescence. We analyze temporal characteristics of NADH fluorescence in a mixture of NADH and lactate dehydrogenase (LDH) as well as in living cell samples. The results show that in both the NADH/LDH mixture and cell samples, a fraction of free NADH and protein-bound components can be identified. The extracted free and bound NADH signals are confirmed by time-resolved measurement of anisotropy decay of NADH fluorescence, based on the fact that free NADH is a small fluorescent molecule with much shorter rotational diffusion time than bound NADH. The ratio of free NADH signal to bound NADH signal is very different between normal and cancer cervical epithelial cells. In addition, the ratio changes significantly when the cell samples are treated with a mitochondrial inhibitor or uncoupler, demonstrating that the method is sensitive to monitor cellular metabolic activity. Finally, we demonstrate that the microviscosity for relatively small molecules such as NADH in cells could be extracted from wavelength- and time-resolved NADH fluorescence of living cell samples.

Nearly half a million children and young adults are affected by traumatic brain injury each year in the United States. Although adequate cerebral blood flow (CBF) is essential to recovery, complications that disrupt blood flow to the brain and exacerbate neurological injury often go undetected because no adequate bedside measure of CBF exists. In this study we validate a depth-resolved, near-infrared spectroscopy (NIRS) technique that provides quantitative CBF measurement despite significant signal contamination from skull and scalp tissue. The respiration rates of eight anesthetized pigs (weight: 16.2±0.5 kg, age: 1 to 2 months old) are modulated to achieve a range of CBF levels. Concomitant CBF measurements are performed with NIRS and CT perfusion. A significant correlation between CBF measurements from the two techniques is demonstrated (r²=0.714, slope=0.92, p<0.001), and the bias between the two techniques is -2.83 mL·min^-1·100 g^-1 (CI_0.95: -19.63 mL·min^-1·100 g^-1-13.9 mL·min^-1·100 g^-1). This study demonstrates that accurate measurements of CBF can be achieved with depth-resolved NIRS despite significant signal contamination from scalp and skull. The ability to measure CBF at the bedside provides a means of detecting, and thereby preventing, secondary ischemia during neurointensive care.

We demonstrate a method to estimate the concentrations of water and lipid in scattering media such as biological tissues with diffuse optical spectra acquired over the range of 900 to 1600 nm. Estimations were performed by fitting the spectra to a model of light propagation in scattering media derived from diffusion theory. To validate the method, spectra were acquired from tissue phantoms consisting of lipid and water emulsions and swine tissues ex vivo with a two-fiber probe.

Intrinsic Raman spectra of biological tissue are distorted by the influences of tissue absorption and scattering, which significantly challenge signal quantification. A combined Raman and spatially resolved reflectance setup is introduced to measure the absorption coefficient µa and the reduced scattering coefficient µ of the tissue, together with the Raman signals. The influence of µa and µ on the resonance Raman signal of -carotene is measured at 1524 cm−1 by tissue phantom measurements and Monte Carlo simulations for µa=0.01 to 10 mm−1 and µ=0.1 to 10 mm−1. Both methods show that the Raman signal drops roughly proportional to 1/µa for µa>0.2 mm−1 in the measurement geometry and that the influence of µ is weaker, but not negligible. Possible correction functions dependent on the elastic diffuse reflectance are investigated to correct the Raman signal for the influence of µa and µ, provided that µa and µ are measured as well. A correction function based on the Monte Carlo simulation of Raman signals is suggested as an alternative. Both approaches strongly reduce the turbidity-induced variation of the Raman signals and allow absolute Raman scattering coefficients to be determined.

We first report on the implementation of a novel narrow-band image-guided Raman endoscopy technique for in vivo diagnosis of gastric dysplasia. High-quality in vivo Raman spectra can be acquired from normal and dysplastic gastric mucosal tissue within 0.5 sec under narrow-band image (NBI) guidance at gastroscopy. Significant differences are observed in in vivo Raman spectra between normal (n=54) and dysplastic (n=18) gastric tissue from 30 gastric patients, particularly in the spectral ranges of 825 to 950, 1000 to 1100, 1250 to 1500, and 1600 to 1800 cm⁻¹, which primarily contain signals related to proteins, nucleic acids, and lipids. The multivariate analysis [i.e., principal components analysis (PCA) and linear discriminant analysis (LDA)], together with the leave-one tissue site-out, cross validation on in vivo gastric Raman spectra yields a diagnostic sensitivity of 94.4% (17/18) and specificity of 96.3% (52/54) for distinction of gastric dysplastic tissue. This study suggests that narrowband image-guided Raman endoscopy associated with PCA-LDA diagnostic algorithms has potential for the noninvasive, in vivo early diagnosis and detection of gastric precancer during clinical gastroscopic examination.

Our aim is to explore the welding capabilities of a thulium (Tm:YAP) laser in modulated and continuous-wave (CW) modes of operation. The Tm:YAP laser system developed for this study includes a Tm:YAP laser resonator, diode laser driver, water chiller, modulation controller unit, and acquisition/control software. Full-thickness incisions on Wistar rat skin were welded by the Tm:YAP laser system at 100 mW and 5 s in both modulated and CW modes of operation (34.66 W/cm²). The skin samples were examined during a 21-day healing period by histology and tensile tests. The results were compared with the samples closed by conventional suture technique. For the laser groups, immediate closure at the surface layers of the incisions was observed. Full closures were observed for both modulated and CW modes of operation at day 4. The tensile forces for both modulated and CW modes of operation were found to be significantly higher than the values found by conventional suture technique. The 1980-nm Tm:YAP laser system operating in both modulated and CW modes maximizes the therapeutic effect while minimizing undesired side effects of laser tissue welding. Hence, it is a potentially important alternative tool to the conventional suturing technique.

Nonablative thermal laser therapy with a 1540-nm laser induces controlled, spatially determined thermal damage, allowing subsequent collagen remodeling while preserving the epidermis. A photorejuvenation effect using nonthermal nonablative stimulation of cells with low energy and narrow band light has been termed photomodulation. Light emitting diodes (LEDs) are narrow band emitters that lead to photomodulation via stimulation of mitochondrial cell organelles. In a previous study, we demonstrated in a human skin explant model that heat shock protein 70 (HSP70) plays a pivotal role in the initiation of skin remodeling after ablative fractional photothermolysis. To test its importance in nonablative laser therapy and photomodulation, the spatio-temporal expression of HSP70 is investigated in response to a 1540-nm laser treatment and six different LED therapies. An Er:glass laser is used with a 1-Hz repetition rate, 30-J/cm² fluence, and a hand piece with a 2-mm spot size. Nonthermal nonablative treatment is performed using two LED (LEDA SCR red light: 635 nm, 40 to 120 W/cm², 40 to 120 J/cm²; LEDA SCR yellow light: 585 nm, 16 to 35 W/cm², 20 to 100 J/cm²; spot size 16×10 cm). Immediate responses as well as responses 1, 3, or 7 days postprocedure are studied; untreated skin explants serve as control. Immunohistochemical investigation (HSP70) is performed in all native, nontreated, and Er:glass laser- or LED-treated samples (n=175). Nonablative laser therapy leads to a clear time-dependent induction of epidermally expressed HSP70, peaking between one to three days post-treatment. In contrast, none of the various LED treatments up-regulated the HSP70 expression in our skin explant model. HSP70 is up-regulated by nonablative but thermal laser devices, but does not seem to play a significant role in the induction of skin remodeling induced by photomodulation.

The use of ultrashort pulse lasers is current in refractive surgery and has recently been extended to corneal grafting (keratoplasty). When performing keratoplasty, however, permanent degradation of the optical properties of the patient's cornea compromises the penetration depth of the laser and the quality of the incisions, therefore causing unwanted secondary effects. Additionally, corneal grafting needs considerably higher penetration depths than refractive surgery. Little data are available about the interaction processes of the femtosecond pulses in the volume of pathological corneas-i.e., in the presence of spherical aberrations and optical scattering. We investigate the influence of the focusing numerical aperture on the laser-tissue interaction. We point out that at low numerical apertures (NAs), tissue damage is produced below and above the focal region. We attribute this phenomenon to nonlinear self-focusing effects. On the other hand, at high NAs, spherical aberrations become significant when focusing at high depths for posterior surgeries, which also limit the cutting efficiency. As high NAs are advisable for reducing unwanted nonlinear effects and ensure accurate cutting, particular attention should be paid to aberration management when developing clinical femtosecond lasers.

This PDF file contains the errata for “JBO Vol. 15 Issue 03 Paper JBO ERR-MAY2010-1” for JBO Vol. 15 Issue 03

This PDF file contains the errata for “JBO Vol. 15 Issue 03 Paper JBO ERR-MAY2010-2” for JBO Vol. 15 Issue 03