Laser Speckle Contrast Imaging (LSCI) is a flexible, easy-to-implement technique for measuring blood flow speeds in-vivo. In order to obtain reliable quantitative data from LSCI the object must remain in the focal plane of the imaging system for the duration of the measurement session. However, since LSCI suffers from inherent frame-to-frame noise, it often requires a moving average filter to produce quantitative results. This frame-to-frame noise also makes the implementation of rapid autofocus system challenging. In this work, we demonstrate an autofocus method and system based on a novel measure of misfocus which serves as an accurate and noise-robust feedback mechanism. This measure of misfocus is shown to enable the localization of best focus with sub-depth-of-field sensitivity, yielding more accurate estimates of blood flow speeds and blood vessel diameters.
(110.6150) Speckle imaging; (110.7348) Wavefront encoding; (170.6480) Spectroscopy, speckle
We propose a new method for phase retrieval that uses partially coherent illumination created by any arbitrary source shape in Köhler geometry. Using a stack of defocused intensity images, we recover not only the phase and amplitude of the sample, but also an estimate of the unknown source shape, which describes the spatial coherence of the illumination. Our algorithm uses a Kalman filtering approach which is fast, accurate and robust to noise. The method is experimentally simple and flexible, so should find use in optical, electron, X-ray and other phase imaging systems which employ partially coherent light. We provide an experimental demonstration in an optical microscope with various condenser apertures.
(100.5070) Phase retrieval; (110.3010) Image reconstruction techniques
The use of fluorescence is ubiquitously found in the detection of immunoreaction; though with good sensitivity, this technique requires labeling as well as other time-consuming steps to perform the measurement. An alternative approach involving liquid crystals (LCs) was proposed, based on the fact that an immunocomplex can disturb the orientation of LCs, leading to an optical texture different from the case when only antigen or antibody exists. This method is label-free, easy to manipulate and low-cost. However, its sensitivity was low for practical usage. In this study, we adopted a high-birefringence liquid crystal (LC) to enhance the sensitivity for the immunodetection. Experiments were performed, targeting at the cancer biomarker CA125. We showed that the larger birefringence (Δn = 0.33 at 20 °C) amplifies the detected signal and, in turn, dramatically improves the detection limit. To avoid signal loss from conventional rinsing steps in immunodetection, CA125 antigen and antibody were reacted before immobilized on substrates. We studied the specific binding events and obtained a detection limit as low as 1 ng/ml. The valid temperature ranges were compared by using the typical single-compound LC 5CB and the high-birefringence LC mixture. We further investigated time dependency of the optical textures and affirmed the capability of LC-based immunodetection in distinguishing between specific and nonspecific antibodies.
(160.3710) Liquid crystals; (170.0170) Medical optics and biotechnology
We correct the omission of the construct and protein purification method in our recent paper [Biomed. Opt. Express 4(12), 2835–2845 (2013)].
(170.0170) Medical optics and biotechnology; (170.1420) Biology; (170.4520) Optical confinement and manipulation; (350.4855) Optical tweezers or optical manipulation
Polarization mode dispersion (PMD) degrades the performance of Jones-matrix-based polarization-sensitive multifunctional optical coherence tomography (JM-OCT). The problem is specially acute for optically buffered JM-OCT, because the long fiber in the optical buffering module induces a large amount of PMD. This paper aims at presenting a method to correct the effect of PMD in JM-OCT. We first mathematically model the PMD in JM-OCT and then derive a method to correct the PMD. This method is a combination of simple hardware modification and subsequent software correction. The hardware modification is introduction of two polarizers which transform the PMD into global complex modulation of Jones matrix. Subsequently, the software correction demodulates the global modulation. The method is validated with an experimentally obtained point spread function with a mirror sample, as well as by in vivo measurement of a human retina.
(110.4500) Optical coherence tomography; (110.5405) Polarimetric imaging; (120.5410) Polarimetry; (170.4500) Optical coherence tomography; (170.4460) Ophthalmic optics and devices; (170.4470) Ophthalmology
Abstract: Measurement of bioluminescent or fluorescent optical reporters with an implanted fiber-optic probe is a promising approach to allow real-time monitoring of molecular and cellular processes in conscious behaving animals. Technically, this approach relies on sensitive light detection due to the relatively limited light signal and inherent light attenuation in scattering tissue. In this paper, we show that specific geometries of lensed fiber probes improve photon collection in turbid tissue such as brain. By employing Monte Carlo simulation and experimental measurement, we demonstrate that hemispherical- and axicon-shaped lensed fibers increase collection efficiency by up to 2-fold when compared with conventional bare fiber. Additionally we provide theoretical evidence that axicon lenses with specific angles improve photon collection over a wider axial range while conserving lateral collection when compared to hemispherical lensed fiber. These findings could guide the development of a minimally-invasive highly sensitive fiber optic-based light signal monitoring technique and may have broad implications such as fiber-based detection used in diffuse optical spectroscopy.
(000.4430) Numerical approximation and analysis; (060.2300) Fiber measurements; (170.3660) Light propagation in tissues; (170.5280) Photon migration; (170.7050) Turbid media
This paper describes an endoscopic-inspired imaging system employing a micro-electromechanical system (MEMS) micromirror scanner to achieve beam scanning for optical coherence tomography (OCT) imaging. Miniaturization of a scanning mirror using MEMS technology can allow a fully functional imaging probe to be contained in a package sufficiently small for utilization in a working channel of a standard gastroesophageal endoscope. This work employs advanced image processing techniques to enhance the images acquired using the MEMS scanner to correct non-idealities in mirror performance. The experimental results demonstrate the effectiveness of the proposed technique.
(230.4685) Optical microelectromechanical devices; (230.0230) Optical devices; (170.2150) Endoscopic imaging; (100.0100) Image processing
To improve the reproducibility of photocoagulation, the ability to quantitatively monitor the thermal change of laser-irradiated retinal tissue is required. Recently, optical coherence tomography has enabled non-invasive and non-contact monitoring of the tissue structural changes during laser irradiation. To further improve the capability of this technique, a method is proposed to measure tissue displacement by simultaneously using Doppler phase shifts and correlation coefficients. The theoretical approach for this method is described, and its performance is experimentally confirmed and evaluated. Finally, lateral and axial displacements in the laser-irradiated retinal tissues of an enucleated porcine eye are observed. The proposed method is found to be useful for further understanding the direct thermal response of laser-irradiated retinal tissue.
(170.4500) Optical coherence tomography; (170.4470) Ophthalmology; (120.6150) Speckle imaging; (170.3340) Laser Doppler velocimetry; (170.5755) Retina scanning
Diagnosis of articular cartilage pathology in the early disease stages using current clinical diagnostic imaging modalities is challenging, particularly because there is often no visible change in the tissue surface and matrix content, such as proteoglycans (PG). In this study, we propose the use of near infrared (NIR) spectroscopy to spatially map PG content in articular cartilage. The relationship between NIR spectra and reference data (PG content) obtained from histology of normal and artificially induced PG-depleted cartilage samples was investigated using principal component (PC) and partial least squares (PLS) regression analyses. Significant correlation was obtained between both data (R2 = 91.40%, p<0.0001). The resulting correlation was used to predict PG content from spectra acquired from whole joint sample, this was then employed to spatially map this component of cartilage across the intact sample. We conclude that NIR spectroscopy is a feasible tool for evaluating cartilage contents and mapping their distribution across mammalian joint.
(170.6510) Spectroscopy, tissue diagnostics; (170.6935) Tissue characterization; (170.3880) Medical and biological imaging
Microcystic macular edema (MME) manifests as small, hyporeflective cystic areas within the retina. For reasons that are still largely unknown, a small proportion of patients with multiple sclerosis (MS) develop MME—predominantly in the inner nuclear layer. These cystoid spaces, denoted pseudocysts, can be imaged using optical coherence tomography (OCT) where they appear as small, discrete, low intensity areas with high contrast to the surrounding tissue. The ability to automatically segment these pseudocysts would enable a more detailed study of MME than has been previously possible. Although larger pseudocysts often appear quite clearly in the OCT images, the multi-frame averaging performed by the Spectralis scanner adds a significant amount of variability to the appearance of smaller pseudocysts. Thus, simple segmentation methods only incorporating intensity information do not perform well. In this work, we propose to use a random forest classifier to classify the MME pixels. An assortment of both intensity and spatial features are used to aid the classification. Using a cross-validation evaluation strategy with manual delineation as ground truth, our method is able to correctly identify 79% of pseudocysts with a precision of 85%. Finally, we constructed a classifier from the output of our algorithm to distinguish clinically identified MME from non-MME subjects yielding an accuracy of 92%.
(100.0100) Image processing; (170.4470) Ophthalmology; (170.4500) Optical coherence tomography
Curing/preventing micrometastasis to lymph nodes (LNs) located outside the surgically resected area is essential for improving the morbidity and mortality associated with breast cancer and head and neck cancer. However, no lymphatic therapy system exists that can deliver drugs to LNs located outside the dissection area. Here, we demonstrate proof of concept for a drug delivery system using MXH10/Mo-lpr/lpr mice that exhibit systemic lymphadenopathy, with some peripheral LNs being as large as 10 mm in diameter. We report that a fluorescent solution injected into the subiliac LN (defined as the upstream LN within the dissection area) was delivered successfully to the proper axillary LN (defined as the downstream LN outside the dissection area) through the lymphatic vessels. Our results suggest that this approach could be used before surgical resection to deliver drugs to downstream LNs outside the dissection area. We anticipate that our methodology could be applied clinically, before surgical resection, to cure/prevent micrometastasis in LNs outside the dissection area, using techniques such as ultrasound-guided internal jugular vein catheterization.
(170.0170) Medical optics and biotechnology; (170.3880) Medical and biological imaging; (170.5380) Physiology
In current clinical practice, the diagnosis of cervical cancer (CC) is mainly through the cervical screening followed by a necessary biopsy, but this method is labor consuming and expensive, and can only detect superficial lesions around the external cervical orifice. In contrast, photoacoustic imaging (PAI) is sensitive to the abnormal angiogenesis deep in the biological tissue, and may be capable for the intact scanning both around the external orifice and in cervical canal. In this paper, we for the first time put forward the photoacoustic diagnosis of CC. A total of 30 in-vitro experiments were carried out in this study, and the obtained depth maximum amplitude projection (DMAP) images were analyzed to evaluate the extent of the angiogenesis for different clinical stages of CC. Stronger absorption from the cervical lesions is observed relative to that of normal tissue. Paired t-test indicates that the difference in mean optical absorption (MOA) between normal tissue and cervical lesion has statistical significance with a confidential coefficient of 0.05. Statistical results also show that the MOAs of the cervical lesions are closely related to the severity of CC. These results imply that PAI may have great utility in the clinical diagnosis of CC.
(170.5120) Photoacoustic imaging; (170.3880) Medical and biological imaging; (170.2150) Endoscopic imaging
We implement an optical tweezers technique to assess the effects of chemical agents on single bacterial cells. As a proof of principle, the viability of a trapped Escherichia coli bacterium is determined by monitoring its flagellar motility in the presence of varying concentrations of ethyl alcohol. We show that the “killing time” of the bacterium can be effectively identified from the correlation statistics of the positional time series recorded from the trap, while direct quantification from the time series or associated power spectra is intractable. Our results, which minimize the lethal effects of bacterial photodamage, are consistent with previous reports of ethanol toxicity that used conventional culture-based methods. This approach can be adapted to study other pairwise combinations of drugs and motile bacteria, especially to measure the response times of single cells with better precision.
(350.4855) Optical tweezers or optical manipulation; (170.4520) Optical confinement and manipulation; (170.3650) Lifetime-based sensing
The stretching stiffness of Red Blood Cells (RBCs) was investigated using a combination of an AC dielectrophoretic apparatus and a single-beam optical tweezer. The experiments were performed at 10 MHz, a frequency high enough to avoid conductivity losses, but below the second turnover point between positive and negative dielectrophoresis. By measuring the geometrical parameters of single healthy human RBCs as a function of the applied voltage, the elastic modulus of RBCs was determined (µ = 1.80 ± 0.5 µN/m) and compared with similar values of the literature got by other techniques. The method is expected to be an easy-to-use, alternative tool to determine the mechano-elastic properties of living cells, and, on this basis, to distinguish healthy and diseased cells
(350.4855) Optical tweezers or optical manipulation; (230.2090) Electro-optical devices; (170.3880) Medical and biological imaging
We propose a robust method for automated computation of heart rate (HR) from digital color video recordings of the human face. In order to extract photoplethysmographic signals, two orthogonal vectors of RGB color space are used. We used a dual tree complex wavelet transform based denoising algorithm to reduce artifacts (e.g. artificial lighting, movement, etc.). Most of the previous work on skin color based HR estimation performed experiments with healthy volunteers and focused to solve motion artifacts. In addition to healthy volunteers we performed experiments with child patients in pediatric intensive care units. In order to investigate the possible factors that affect the non-contact HR monitoring in a clinical environment, we studied the relation between hemoglobin levels and HR estimation errors. Low hemoglobin causes underestimation of HR. Nevertheless, we conclude that our method can provide acceptable accuracy to estimate mean HR of patients in a clinical environment, where the measurements can be performed remotely. In addition to mean heart rate estimation, we performed experiments to estimate oxygen saturation. We observed strong correlations between our SpO2 estimations and the commercial oximeter readings
(170.3880) Medical and biological imaging; (100.7410) Wavelets; (100.2960) Image analysis
Light scattering in biological tissue significantly limits the accessible depth for localized optical interrogation and deep-tissue optical imaging. This challenge can be overcome by exploiting the time-reversal property of optical phase conjugation (OPC) to reverse multiple scattering events or suppress turbidity. However, in living tissue, scatterers are highly movable and the movement can disrupt time-reversal symmetry when there is a latency in the OPC playback. In this paper, we show that the motion-induced degradation of the OPC turbidity-suppression effect through a dynamic scattering medium shares the same decorrelation time constant as that determined from speckle intensity autocorrelation – a popular conventional measure of scatterer movement. We investigated this decorrelation characteristic time through a 1.5-mm-thick dorsal skin flap of a living mouse and found that it ranges from 50 ms to 2.5 s depending on the level of immobilization. This study provides information on relevant time scales for applying OPC to living tissues.
(070.5040) Phase conjugation; (170.7050) Turbid media; (190.5040) Phase conjugation; (090.1995) Digital holography; (110.1080) Active or adaptive optics
Multimodal spectral imaging (MSI) based on auto-fluorescence imaging and Raman micro-spectroscopy was used to detect basal cell carcinoma (BCC) in tissue specimens excised during Mohs micrographic surgery. In this study, the MSI algorithm was optimized to maximize the diagnosis accuracy while minimizing the number of Raman spectra: the segmentation of the auto-fluorescence images was optimized according to the type of BCC, sampling points for Raman spectroscopy were generated based on auto-fluorescence intensity variance and segment area, additional Raman spectra were acquired when performance of the segmentation algorithm was sub-optimal. The results indicate that accurate diagnosis can be achieved with a sampling density of ~2,000 Raman spectra/cm2, based on sampling points generated by the MSI algorithms. The key benefit of MSI is that diagnosis of BCC is obtained based on intrinsic chemical contrast of the tissue, within time scales similar to frozen-section histopathology, but without requiring laborious sample preparation and subjective interpretation of stained frozen-sections.
(170.0170) Medical optics and biotechnology; (170.5660) Raman spectroscopy; (170.4580) Optical diagnostics for medicine; (170.1870) Dermatology
Tissue optical and mechanical properties are correlated to tissue pathologic changes. This manuscript describes a dual-mode ultrasound modulated optical imaging system capable of sensing local optical and mechanical properties in reflection geometry. The optical characterisation was achieved by the acoustic radiation force assisted ultrasound modulated optical tomography (ARF-UOT) with laser speckle contrast detection. Shear waves generated by the ARF were also tracked optically by the same system and the shear wave speed was used for the elasticity measurement. Tissue mimicking phantoms with multiple inclusions buried at 11 mm depth were experimentally scanned with the dual-mode system. The inclusions, with higher optical absorption and/or higher stiffness than background, were identified based on the dual results and their stiffnesses were quantified. The system characterises both optical and mechanical properties of the inclusions compared with the ARF-UOT or the elasticity measurement alone. Moreover, by detecting the backward scattered light in reflection detection geometry, the system is more suitable for clinical applications compared with transmission geometry.
(170.1065) Acousto-optics; (110.0113) Imaging through turbid media
A micro flow cytometer with a single step 3D hydrodynamic flow focusing has been developed. The proposed design is capable to create a single-file particle stream that is self-aligned with an integrated optical fiber-based detection system, regardless of the flow rate ratio between the focusing and core liquids. The design approach provides the ability to adjust the stream size while keeping the position of the focused stream centered with respect to the focusing channel. The device has been fabricated by direct micro milling of PMMA sheets. Experimental validation of the hydrodynamic sheath focusing effect has been presented and sample stream with tuneable size from about 18 to 50 μm was measured. Flow cytometry measurements have been performed by using 10-23 μm fluorescent particles. From the analysis of the signals collected at each transit event we can confirm that the device was capable to align and measure microparticles with a good coefficient of variance.
(230.3990) Micro-optical devices; (300.2530) Fluorescence, laser-induced; (120.5820) Scattering measurements
Gas assisted laser machining of materials is a common practice in the manufacturing industry. Advantages in using gas assistance include reducing the likelihood of flare-ups in flammable materials and clearing away ablated material in the cutting path. Current surgical procedures and research do not take advantage of this and in the case for resecting osseous tissue, gas assisted ablation can help minimize charring and clear away debris from the surgical site. In the context of neurosurgery, the objective is to cut through osseous tissue without damaging the underlying neural structures. Different inert gas flow rates used in laser machining could cause deformations in compliant materials. Complications may arise during surgical procedures if the dura and spinal cord are damaged by these deformations. We present preliminary spinal deformation findings for various gas flow rates by using optical coherence tomography to measure the depression depth at the site of gas delivery.
(110.0110) Imaging systems; (110.4500) Optical coherence tomography; (140.0140) Lasers and laser optics; (140.3460) Lasers; (170.0170) Medical optics and biotechnology; (170.1020) Ablation of tissue; (170.3880) Medical and biological imaging
Femtosecond laser pulses can be used to perform very precise cutting of material, including biological samples from subcellular organelles to large areas of bone, through plasma-mediated ablation. The use of a kilohertz regenerative amplifier is usually needed to obtain the pulse energy required for ablation. This work investigates a 5 megahertz compact fiber laser for near-video rate imaging and ablation in bone. After optimization of ablation efficiency and reduction in autofluorescence, the system is demonstrated for the in vivo study of bone regeneration. Image-guided creation of a bone defect and longitudinal evaluation of cellular injury response in the defect provides insight into the bone regeneration process.
(170.1020) Ablation of tissue; (170.2520) Fluorescence microscopy; (060.4370) Nonlinear optics, fibers; (180.4315) Nonlinear microscopy
We demonstrate a novel non-contact method: acoustic radiation force impulse microscopy via photoacoustic detection (PA-ARFI), capable of probing cell mechanics. A 30 MHz lithium niobate ultrasound transducer is utilized for both detection of phatoacoustic signals and generation of acoustic radiation force. To track cell membrane displacements by acoustic radiation force, functionalized single-walled carbon nanotubes are attached to cell membrane. Using the developed microscopy evaluated with agar phantoms, the mechanics of highly- and weakly-metastatic breast cancer cells are quantified. These results clearly show that the PA-ARFI microscopy may serve as a novel tool to probe mechanics of single breast cancer cells.
(170.7170) Ultrasound; (170.5120) Photoacoustic imaging; (170.0180) Microscopy
Compared to deep brain electrical stimulation, which has been applied to treating pathological brain diseases, little work has been done on the effect of deep brain light stimulation. A fiber-coupled laser stimulator at 840 nm wavelength and 130 Hz pulse repetition rate is developed in this work for deep brain light stimulation in a rat model. Concentration changes in glutamate and dopamine in the striatum are observed using a microdialysis probe when the subthalamic nucleus (STN) is stimulated at various optical power levels. Experimental results show that light stimulation causes the concentration of glutamate to decrease while that of dopamine is increased. This suggests that deep brain light stimulation of the STN is a promising therapeutic strategy for dopamine-related diseases such as Parkinson’s disease. The stimulator developed for this work is useful for deep brain light stimulation in biomedical research.
(170.5180) Photodynamic therapy; (170.3660) Light propagation in tissues; (170.3890) Medical optics instrumentation; (170.1420) Biology; (170.1610) Clinical applications; (170.0170) Medical optics and biotechnology
Diffuse optical tomography for medical applications can require probes with small dimensions involving short source-detector separations. Even though this configuration is seen at first as a constraint due to the challenge of depth sensitivity, we show here that it can potentially be an asset for spatial resolution in depth. By comparing two fiber optic probes on a test object, we first show with simulations that short source-detector separations improve the spatial resolution down to a limit depth. We then confirm these results in an experimental study with a state-of-the-art setup involving a fast-gated single-photon avalanche diode allowing maximum depth sensitivity. We conclude that short source-detector separations are an option to consider for the design of probes so as to improve image quality for diffuse optical tomography in reflectance.
(170.3880) Medical and biological imaging; (170.6960) Tomography; (170.6920) Time-resolved imaging; (170.3010) Image reconstruction techniques; (170.7050) Turbid media; (230.5160) Photodetectors
Hemodynamics plays a critical role in the development of atherosclerosis, specifically in regions of curved vasculature such as bifurcations exhibiting irregular blood flow profiles. Carotid atherosclerotic disease can be intervened by stent implantation, but this may result in greater alterations to local blood flow and consequently further complications. This study demonstrates the use of a variant of Doppler optical coherence tomography (DOCT) known as split spectrum DOCT (ssDOCT) to evaluate hemodynamic patterns both before and after stent implantation in the bifurcation junction in the internal carotid artery (ICA). Computational fluid dynamics (CFD) models were constructed to simulate blood velocity profiles and compared to the findings achieved through ssDOCT images. Both methods demonstrated noticeable alterations in hemodynamic patterns following stent implantation, with features such as slow velocity regions at the neck of the bifurcation and recirculation zones at the stent struts. Strong correlation between CFD models and ssDOCT images demonstrate the potential of ssDOCT imaging in the optimization of stent implantation in the clinical setting.
(170.4500) Optical coherence tomography; (170.3880) Medical and biological imaging; (120.5050) Phase measurement