Performance metrics of an optical spectral imaging system for intra-operative assessment of breast tumor margins.

As many as 20-70% of patients undergoing breast conserving surgery require repeat surgeries due to a close or positive surgical margin diagnosed post-operatively [1]. Currently there are no widely accepted tools for intra-operative margin assessment which is a significant unmet clinical need. Our group has developed a first-generation optical visible spectral imaging platform to image the molecular composition of breast tumor margins and has tested it clinically in 48 patients in a previously published study [2]. The goal of this paper is to report on the performance metrics of the system and compare it to clinical criteria for intra-operative tumor margin assessment. The system was found to have an average signal to noise ratio (SNR) >100 and <15% error in the extraction of optical properties indicating that there is sufficient SNR to leverage the differences in optical properties between negative and close/positive margins. The probe had a sensing depth of 0.5-2.2 mm over the wavelength range of 450-600 nm which is consistent with the pathologic criterion for clear margins of 0-2 mm. There was <1% cross-talk between adjacent channels of the multi-channel probe which shows that multiple sites can be measured simultaneously with negligible cross-talk between adjacent sites. Lastly, the system and measurement procedure were found to be reproducible when evaluated with repeated measures, with a low coefficient of variation (<0.11). The only aspect of the system not optimized for intra-operative use was the imaging time. The manuscript includes a discussion of how the speed of the system can be improved to work within the time constraints of an intra-operative setting.

Probing polarization and dielectric function of molecules with higher order harmonics in scattering-near-field scanning optical microscopy

The idealized system of an atomically flat metallic surface [highly oriented pyrolytic graphite (HOPG)] and an organic monolayer (porphyrin) was used to determine whether the dielectric function and associated properties of thin films can be accessed with scanning-near-field scanning optical microscopy (s-NSOM). Here, we demonstrate the use of harmonics up to fourth order and the polarization dependence of incident light to probe dielectric properties on idealized samples of monolayers of organic molecules on atomically smooth substrates. An analytical treatment of light/sample interaction using the s-NSOM tip was developed in order to quantify the dielectric properties. The theoretical analysis and numerical modeling, as well as experimental data, demonstrate that higher order harmonic scattering can be used to extract the dielectric properties of materials with tens of nanometer spatial resolution. To date, the third harmonic provides the best lateral resolution (∼50 nm) and dielectric constant contrast for a porphyrin film on HOPG. © 2009 American Institute of Physics.

Intraoperative spectral domain optical coherence tomography for vitreoretinal surgery.

We demonstrate in vivo human retinal imaging using an intraoperative microscope-mounted optical coherence tomography system (MMOCT). Our optomechanical design adapts an Oculus Binocular Indirect Ophthalmo Microscope (BIOM3), suspended from a Leica ophthalmic surgical microscope, with spectral domain optical coherence tomography (SD-OCT) scanning and relay optics. The MMOCT enables wide-field noncontact real-time cross-sectional imaging of retinal structure, allowing for SD-OCT augmented intrasurgical microscopy for intraocular visualization. We experimentally quantify the axial and lateral resolution of the MMOCT and demonstrate fundus imaging at a 20Hz frame rate.

Fiber-optic interferometric two-dimensional scattering-measurement system.

We present a fiber-optic interferometric system for measuring depth-resolved scattering in two angular dimensions using Fourier-domain low-coherence interferometry. The system is a unique hybrid of the Michelson and Sagnac interferometer topologies. The collection arm of the interferometer is scanned in two dimensions to detect angular scattering from the sample, which can then be analyzed to determine the structure of the scatterers. A key feature of the system is the full control of polarization of both the illumination and the collection fields, allowing for polarization-sensitive detection, which is essential for two-dimensional angular measurements. System performance is demonstrated using a double-layer microsphere phantom. Experimental data from samples with different sizes and acquired with different polarizations show excellent agreement with Mie theory, producing structural measurements with subwavelength accuracy.

A low-cost, portable, and quantitative spectral imaging system for application to biological tissues.

The ability of diffuse reflectance spectroscopy to extract quantitative biological composition of tissues has been used to discern tissue types in both pre-clinical and clinical cancer studies. Typically, diffuse reflectance spectroscopy systems are designed for single-point measurements. Clinically, an imaging system would provide valuable spatial information on tissue composition. While it is feasible to build a multiplexed fiber-optic probe based spectral imaging system, these systems suffer from drawbacks with respect to cost and size. To address these we developed a compact and low cost system using a broadband light source with an 8-slot filter wheel for illumination and silicon photodiodes for detection. The spectral imaging system was tested on a set of tissue mimicking liquid phantoms which yielded an optical property extraction accuracy of 6.40 +/- 7.78% for the absorption coefficient (micro(a)) and 11.37 +/- 19.62% for the wavelength-averaged reduced scattering coefficient (micro(s)').

Generalized sampling using a compound-eye imaging system for multi-dimensional object acquisition.

In this paper, we propose generalized sampling approaches for measuring a multi-dimensional object using a compact compound-eye imaging system called thin observation module by bound optics (TOMBO). This paper shows the proposed system model, physical examples, and simulations to verify TOMBO imaging using generalized sampling. In the system, an object is modulated and multiplied by a weight distribution with physical coding, and the coded optical signal is integrated on to a detector array. A numerical estimation algorithm employing a sparsity constraint is used for object reconstruction.

3D refraction correction and extraction of clinical parameters from spectral domain optical coherence tomography of the cornea.

Capable of three-dimensional imaging of the cornea with micrometer-scale resolution, spectral domain-optical coherence tomography (SDOCT) offers potential advantages over Placido ring and Scheimpflug photography based systems for accurate extraction of quantitative keratometric parameters. In this work, an SDOCT scanning protocol and motion correction algorithm were implemented to minimize the effects of patient motion during data acquisition. Procedures are described for correction of image data artifacts resulting from 3D refraction of SDOCT light in the cornea and from non-idealities of the scanning system geometry performed as a pre-requisite for accurate parameter extraction. Zernike polynomial 3D reconstruction and a recursive half searching algorithm (RHSA) were implemented to extract clinical keratometric parameters including anterior and posterior radii of curvature, central cornea optical power, central corneal thickness, and thickness maps of the cornea. Accuracy and repeatability of the extracted parameters obtained using a commercial 859nm SDOCT retinal imaging system with a corneal adapter were assessed using a rigid gas permeable (RGP) contact lens as a phantom target. Extraction of these parameters was performed in vivo in 3 patients and compared to commercial Placido topography and Scheimpflug photography systems. The repeatability of SDOCT central corneal power measured in vivo was 0.18 Diopters, and the difference observed between the systems averaged 0.1 Diopters between SDOCT and Scheimpflug photography, and 0.6 Diopters between SDOCT and Placido topography.

Towards a field-compatible optical spectroscopic device for cervical cancer screening in resource-limited settings: effects of calibration and pressure.

Quantitative optical spectroscopy has the potential to provide an effective low cost, and portable solution for cervical pre-cancer screening in resource-limited communities. However, clinical studies to validate the use of this technology in resource-limited settings require low power consumption and good quality control that is minimally influenced by the operator or variable environmental conditions in the field. The goal of this study was to evaluate the effects of two sources of potential error: calibration and pressure on the extraction of absorption and scattering properties of normal cervical tissues in a resource-limited setting in Leogane, Haiti. Our results show that self-calibrated measurements improved scattering measurements through real-time correction of system drift, in addition to minimizing the time required for post-calibration. Variations in pressure (tested without the potential confounding effects of calibration error) caused local changes in vasculature and scatterer density that significantly impacted the tissue absorption and scattering properties Future spectroscopic systems intended for clinical use, particularly where operator training is not viable and environmental conditions unpredictable, should incorporate a real-time self-calibration channel and collect diffuse reflectance spectra at a consistent pressure to maximize data integrity.

Portable, Fiber-Based, Diffuse Reflection Spectroscopy (DRS) Systems for Estimating Tissue Optical Properties.

Steady-state diffuse reflection spectroscopy is a well-studied optical technique that can provide a noninvasive and quantitative method for characterizing the absorption and scattering properties of biological tissues. Here, we compare three fiber-based diffuse reflection spectroscopy systems that were assembled to create a light-weight, portable, and robust optical spectrometer that could be easily translated for repeated and reliable use in mobile settings. The three systems were built using a broadband light source and a compact, commercially available spectrograph. We tested two different light sources and two spectrographs (manufactured by two different vendors). The assembled systems were characterized by their signal-to-noise ratios, the source-intensity drifts, and detector linearity. We quantified the performance of these instruments in extracting optical properties from diffuse reflectance spectra in tissue-mimicking liquid phantoms with well-controlled optical absorption and scattering coefficients. We show that all assembled systems were able to extract the optical absorption and scattering properties with errors less than 10%, while providing greater than ten-fold decrease in footprint and cost (relative to a previously well-characterized and widely used commercial system). Finally, we demonstrate the use of these small systems to measure optical biomarkers in vivo in a small-animal model cancer therapy study. We show that optical measurements from the simple portable system provide estimates of tumor oxygen saturation similar to those detected using the commercial system in murine tumor models of head and neck cancer.

Waveguide QED: Many-body bound-state effects in coherent and Fock-state scattering from a two-level system

Strong coupling between a two-level system (TLS) and bosonic modes produces dramatic quantum optics effects. We consider a one-dimensional continuum of bosons coupled to a single localized TLS, a system which may be realized in a variety of plasmonic, photonic, or electronic contexts. We present the exact many-body scattering eigenstate obtained by imposing open boundary conditions. Multiphoton bound states appear in the scattering of two or more photons due to the coupling between the photons and the TLS. Such bound states are shown to have a large effect on scattering of both Fock- and coherent-state wave packets, especially in the intermediate coupling-strength regime. We compare the statistics of the transmitted light with a coherent state having the same mean photon number: as the interaction strength increases, the one-photon probability is suppressed rapidly, and the two- and three-photon probabilities are greatly enhanced due to the many-body bound states. This results in non-Poissonian light. © 2010 The American Physical Society.

Live Imaging of Host-Parasite Interactions in a Zebrafish Infection Model Reveals Cryptococcal Determinants of Virulence and Central Nervous System Invasion.

UNLABELLED: The human fungal pathogen Cryptococcus neoformans is capable of infecting a broad range of hosts, from invertebrates like amoebas and nematodes to standard vertebrate models such as mice and rabbits. Here we have taken advantage of a zebrafish model to investigate host-pathogen interactions of Cryptococcus with the zebrafish innate immune system, which shares a highly conserved framework with that of mammals. Through live-imaging observations and genetic knockdown, we establish that macrophages are the primary immune cells responsible for responding to and containing acute cryptococcal infections. By interrogating survival and cryptococcal burden following infection with a panel of Cryptococcus mutants, we find that virulence factors initially identified as important in causing disease in mice are also necessary for pathogenesis in zebrafish larvae. Live imaging of the cranial blood vessels of infected larvae reveals that C. neoformans is able to penetrate the zebrafish brain following intravenous infection. By studying a C. neoformans FNX1 gene mutant, we find that blood-brain barrier invasion is dependent on a known cryptococcal invasion-promoting pathway previously identified in a murine model of central nervous system invasion. The zebrafish-C. neoformans platform provides a visually and genetically accessible vertebrate model system for cryptococcal pathogenesis with many of the advantages of small invertebrates. This model is well suited for higher-throughput screening of mutants, mechanistic dissection of cryptococcal pathogenesis in live animals, and use in the evaluation of therapeutic agents. IMPORTANCE: Cryptococcus neoformans is an important opportunistic pathogen that is estimated to be responsible for more than 600,000 deaths worldwide annually. Existing mammalian models of cryptococcal pathogenesis are costly, and the analysis of important pathogenic processes such as meningitis is laborious and remains a challenge to visualize. Conversely, although invertebrate models of cryptococcal infection allow high-throughput assays, they fail to replicate the anatomical complexity found in vertebrates and, specifically, cryptococcal stages of disease. Here we have utilized larval zebrafish as a platform that overcomes many of these limitations. We demonstrate that the pathogenesis of C. neoformans infection in zebrafish involves factors identical to those in mammalian and invertebrate infections. We then utilize the live-imaging capacity of zebrafish larvae to follow the progression of cryptococcal infection in real time and establish a relevant model of the critical central nervous system infection phase of disease in a nonmammalian model.

Optical-CT 3D Dosimetry Using Fresnel Lenses with Minimal Refractive-Index Matching Fluid.

Telecentric optical computed tomography (optical-CT) is a state-of-the-art method for visualizing and quantifying 3-dimensional dose distributions in radiochromic dosimeters. In this work a prototype telecentric system (DFOS-Duke Fresnel Optical-CT Scanner) is evaluated which incorporates two substantial design changes: the use of Fresnel lenses (reducing lens costs from $10-30K t0 $1-3K) and the use of a 'solid tank' (which reduces noise, and the volume of refractively matched fluid from 1 ltr to 10 cc). The efficacy of DFOS was evaluated by direct comparison against commissioned scanners in our lab. Measured dose distributions from all systems were compared against the predicted dose distributions from a commissioned treatment planning system (TPS). Three treatment plans were investigated including a simple four-field box treatment, a multiple small field delivery, and a complex IMRT treatment. Dosimeters were imaged within 2 h post irradiation, using consistent scanning techniques (360 projections acquired at 1 degree intervals, reconstruction at 2mm). DFOS efficacy was evaluated through inspection of dose line-profiles, and 2D and 3D dose and gamma maps. DFOS/TPS gamma pass rates with 3%/3mm dose difference/distance-to-agreement criteria ranged from 89.3% to 92.2%, compared to from 95.6% to 99.0% obtained with the commissioned system. The 3D gamma pass rate between the commissioned system and DFOS was 98.2%. The typical noise rates in DFOS reconstructions were up to 3%, compared to under 2% for the commissioned system. In conclusion, while the introduction of a solid tank proved advantageous with regards to cost and convenience, further work is required to improve the image quality and dose reconstruction accuracy of the new DFOS optical-CT system.