Preclinical Evaluation of Spatial Frequency Domain-Enabled Wide-Field Quantitative Imaging for Enhanced Glioma Resection

5-Aminolevelunic acid-induced protoporphyrin IX (PpIX) fluorescence-guided resection (FGR) enables maximum safe resection of glioma by providing real-time tumor contrast. However, the subjective visual assessment and the variable intrinsic optical attenuation of tissue limit this technique to reliably delineating only high-grade tumors that display strong fluorescence. We have previously shown, using a fiber-optic probe, that quantitative assessment using noninvasive point spectroscopic measurements of the absolute PpIX concentration in tissue further improves the accuracy of FGR, extending it to surgically curable low-grade glioma. More recently, we have shown that implementing spatial frequency domain imaging with a fluorescent-light transport model enables recovery of two-dimensional images of [PpIX], alleviating the need for time-consuming point sampling of the brain surface. We present first results of this technique modified for in vivo imaging on an RG2 rat brain tumor model. Despite the moderate errors in retrieving the absorption and reduced scattering coefficients in the subdiffusive regime of 14% and 19%, respectively, the recovered [PpIX] maps agree within 10% of the point [PpIX] values measured by the fiber-optic probe, validating its potential as an extension or an alternative to point sampling during glioma resection

Polymer-Based Microfluidic Devices for High Throughput Single Molecule Detection: Applications in Biological and Drug Discovery

The realization of high throughput sample processing has become a primary ambition in many research applications with an example being high throughput screening (HTS), which represents the first step in the drug discovery pipeline. Microfluidics is a viable platform for parallel processing of biochemical reactions to increase data production rates due to its ability to generate fluidic networks with a high number of processors over small footprints suitable for optical imaging. Single-molecule detection (SMD) is another technology which has emerged to facilitate the realization of high throughput data processing afforded by its ability to eliminate sample processing steps and generate results with high statistical accuracy. A combination of microfluidics and SMD with wide-field fluorescence detection provides the ability to monitor biochemical reactions in a high throughput format with single-molecule sensitivity. In this dissertation, the integration of these techniques was presented for HTS applications in drug discovery. An ultra-sensitive fluorescence detection system with a wide field-of-view (FoV) was constructed to transduce fluorescence signatures from single chromophores that were electrokinetically transported through a series of tightly packed fluidic channels poised on poly(methylmethacrylate), PMMA and contained within the FoV of a laser detection system. The system was used to monitor biochemical reactions at the single-molecule level in a continuous-flow format. Enhancement in sampling-throughput was demonstrated by constructing a high density fluidic network for parallel analysis of multiple biochemical assays. In another development, the ability to enhance single-molecule sensitivity in a flow-based biochemical assay was investigated using a novel cyclic olefin copolymer (COC) planar waveguide embedded in PMMA and situated orthogonal to multiple fluidic channels. This design allowed for fluorescence detection from multiple fluidic channels using evanescent excitation and a wide FoV fluorescence detection system for parallel readout. Results from these technologies were presented as well as their applications in drug discovery for increasing data production rates and quality. An approach toward monitoring the efficacy of therapeutic agents, which is important in clinical evaluation of drug potency in the drug discovery process, was also considered, by designing a microfluidic system with integrated conductivity sensor for label-free enumeration of isolated tumor cells from clinical samples

Imaging cytometry technology for environmental and biomedical applications

Early detection of microorganisms in environmental and biomedical applications is critical for the effective response to potential pathogenic treats. Most traditional methods and instrumentation for the analysis of these samples are becoming obsolete due to the fact that they are time-consuming and have long response times. Modern solutions are limited to high-end centralized facilities and specialized trained personnel, given their high-cost and complexity. There is thus a clear need to develop and introduce low-cost, easy to use, high-performance devices capable of rapidly identifying and quantifying pathogenic microorganisms in environmental and biomedical samples. The work behind this thesis was devoted to the design, development and validation in relevant industrial environments of two image cytometry devices capable of characterizing biological and industrial samples in terms of their microorganism content. Bringing a potentially high-impact solution to the current industry need. The first technology, defined within the context of this thesis as Fourier image cytometry, is an optical device capable of increasing the sample volume tested when compared to traditional state-of-the-art counterparts. By evaluating the sample in the Fourier domain, the device is capable of measuring characteristics of particulate within a sample volume larger than other imaging systems. The result is an enhancement of both field of view (FOV) and depth of field (DOF) of the target sample. Furthermore, the implementation of the Fourier image cytometer in this thesis is a portable and compact device comprising of low-cost optics and electronics. The design of the entire device was performed with the objective to minimize the cost and maximize the capabilities. This was possible mainly due to the recent advances in image sensor technologies that simplify the device’s optics. In our Fourier imaging cytometry, LED light sources and conventional achromatic optical lenses are at the basis of device’s optics as opposed to high-end lasers or optical microscopes. For the detection scheme, a CMOS image sensor was used. After optimizing the prototype and going through rigorous validation in a laboratory, the Fourier image cytometry introduced in this thesis was validated in two relevant industrial environments. The device was tested using real environmental samples. In the first industrial validation, it was used for the microorganism’s identification and quantification in water coming from cooling towers. The second industrial validation used a further optimized implementation of the cytometer to analyze fresh and marine waters for their microorganism population, specifically phytoplankton within the context of ballast water and ballast water treatment systems. The second image cytometry designed, developed and implemented within the scope of this thesis, focused on detection of microorganisms spread over surfaces. Following the motivation for low-cost compact devices, a Surface cytometer was designed. The Surface cytometer is an optical device capable of quantifying bacterial population over a surface of over 300 mm2. The device was completely autonomous thanks to the integration of a single-board computer within its design. The light source and detection scheme continued to be LED source and CMOS sensor detection. Similarly to the validation process of the Fourier cytometer, the Surface cytometer was tested in controlled samples in a laboratory environment, before it was put to test on a biomedical application for bacterial growth monitoring and compared to standard devices of measurement of optical density, used today in the industry. In summary, in this thesis we present two novel image cytometers and three clear industrial applications in which the devices were validated. This clearly indicates the potential of image cytometry as an effective low cost and portable tool for the analysis of microorganisms in the environmental and biomedical sectorLa detección temprana de microorganismos en aplicaciones ambientales y biomédicas es crítica para la respuesta efectiva a posibles amenazas patogénicas. La mayoría de los métodos e instrumentos tradicionales para este tipo de análisis están casi obsoletas, debido a los esfuerzos que requieren y sus largos tiempos de respuesta. Las soluciones modernas se limitan a instalaciones centralizadas de alta gama y personal especializado, esto debido a su alto coste y complejidad. Existe una clara necesidad de desarrollar e introducir dispositivos de bajo costo, fáciles de usar y de alto rendimiento capaces de identificar y cuantificar rápidamente microorganismos patogénicos en muestras ambientales y biomédicas. El trabajo detrás de esta tesis se dedicó al diseño, desarrollo y validación en entornos industriales relevantes de dos dispositivos de citometría de imagen. La primera tecnología, definida como citometría de imagen de Fourier, es un dispositivo óptico capaz de aumentar el volumen de muestra capturado en comparación con las tecnologías tradicionales y el estado del arte. Al evaluar la muestra en el dominio de Fourier, el sistema es capaz de medir las características de las partículas dentro de un volumen de muestra mayor que los sistemas de imágenes comparativos. El sistema resultante mejora tanto el campo de visión (FOV, por sus siglas en inglés) como la profundidad de campo (DOF, por sus siglas en inglés) de la muestra. Además, la implementación del citómetro de imagen de Fourier en esta tesis es un dispositivo compacto y portátil compuesto por componentes ópticos y electrónicos de bajo coste. El diseño de todo el sistema se realizó con el objetivo de minimizar el coste del sistema y maximizar sus prestaciones. Esto fue posible principalmente debido a los recientes avances en las tecnologías de sensores de imagen que nos permitieron simplificar la óptica del dispositivo. En esta implementación de la citometría de imagen de Fourier, fuentes de luz LED y las lentes ópticas acromáticas convencionales comprenden la óptica del sistema en lugar de láseres de alta gama u objetivos de microscopios ópticos. Para el esquema de detección se utilizó un sensor de imagen CMOS. La citometría de imagen de Fourier presentada en esta tesis también fue validada en dos entornos industriales relevantes. El sistema se probó utilizando muestras ambientales reales. En la primera validación industrial, el sistema se utilizó para la identificación y cuantificación del microorganismo en el agua proveniente de torres de refrigeración. En la segunda validación industrial se analizaron aguas dulces y marinas, y su población de microorganismos, específicamente la cuantificación de phytoplankton en el contexto de sistemas de tratamiento de aguas de lastre. La segunda citometría de imagen diseñada, desarrollada e implementada dentro del alcance de esta tesis se centró en la detección de microorganismos sobre superficies. Siguiendo la motivación de los dispositivos compactos de bajo costo, se diseñó un citómetro de superficie. El citómetro de superficie es un dispositivo óptico capaz de cuantificar la población bacteriana en una superficie de más de 300 mm2. El dispositivo es completamente autónomo gracias a la integración de una computadora de placa única dentro de su diseño. La fuente de luz y el esquema de detección continuaron siendo LED y sensor CMOS. De manera similar al proceso de validación del citómetro de Fourier, el citómetro de superficie se probó en muestras controladas en un entorno de laboratorio, antes de someterse a prueba en una aplicación biomédica para el monitoreo del crecimiento bacteriano y se comparó con los sistemas estándar de medición de densidad óptica, utilizados hoy en día. en la industria. En resumen, en esta tesis presentamos dos nuevas tecnologías de citometría, junto con dos dispositivos de alto rendimiento y tres aplicaciones industriale

Methods for Focal Plane Array Resolution Estimation Using Random Laser Speckle in Non-paraxial Geometries

The infrared (IR) imaging community has a need for direct IR detector evaluation due to the continued demand for small pixel pitch detectors, the emergence of strained-layer-super-lattice devices, and the associated lateral carrier diffusion issues. Conventional laser speckle-based modulation transfer function (MTF) estimation is dependent on Fresnel propagation and a wide-sense-stationary input random process, limiting the use of this approach for lambda (wavelength)-scale IR devices. This dissertation develops two alternative methodologies for speckle-based resolution evaluation of IR focal plane arrays (FPAs). Both techniques are formulated using Rayleigh-Sommerfield electric field propagation, making them valid in the non-paraxial geometries dictated for resolution estimation of lambda-scale devices. The generalized FPA MTF estimation approach numerically evaluates Rayleigh-Sommerfeld speckle irradiance autocorrelation functions (ACFs) to indirectly compute the power spectral density (PSD) of a non-wide-sense-stationary (WSS) speckle irradiance random process. The experimental error incurred by making WSS assumptions regarding the associated laser speckle random process is quantified utilizing the Wigner distribution function. This method is experimentally demonstrated on a lambda-scale longwave IR FPA, showing a 27% spatial frequency range improvement over established estimation methodology. Additionally, a resolution estimation approach, which utilizes an iterative maximum likelihood estimation approach and speckle irradiance ACFs to solve for a system impulse response, is developed and demonstrated with simulated speckle imagery

Validation of a Confocal Light Sheet Microscope using Push Broom Translation for Biomedical Applications

There exists a need for research of optical methods capable of image cytometry suitable for point-of-care technology. To propose am optical approach with no moving parts for simplification of mechanical components for the further development of the technology to the poin-of-care, a linear sensor with push broom translation method. Push broom translation is a method of moving objects by the sensor for an extended field of view. A polydimethylsiloxane (PDMS) microfluidic chamber with a syringe pump was used to deliver objects by the sensor. The volumetric rate of the pump was correlated to the integration time of the sensor to ensure images were realistically being formed, termed aspect ratio. An electro-chemical microfluidic system was then also investigated, redox-magnetohydrodynamics (R-MHD), to eliminate the mechanical syringe pump which showed deviations in linear speeds at the specimen plane. To image with adequate signal to background ratio within the deep chamber of the R-MHD device, an epitaxial light sheet confocal microscope (e-LSCM) was used to improve axial resolution. The linear sensor, having small pixels, blocked out-of-plane light while eliminating the need for a mechanical aperture which is used for traditional point-scanning confocal microscopy. The particular linear sensor used has binning modes that were used to vary the axial resolution by increasing the sensor aperture. This approach was validated by using a mirror translated in the axial direction and measuring remitted light intensity. The resulting curve estimated the real axial resolution of the microscope, which compared favorably to theoretical values. The R-MHD and the e-LSCM were then synchronized to perform continuous imaging of fluorescent microspheres and cells in suspension. This study combines epitaxial light sheet confocal microscopy and electro-chemical microfluidics as a robust approach which could be used in future point-of-care image cytometry applications

NASA patent abstracts bibliography: A continuing bibliography. Section 1: Abstracts (supplement 43)

Abstracts are provided for 128 patents and patent applications entered into the NASA scientific and technical information system during the period Jan. 1993 through Jun. 1993. Each entry consists of a citation, an abstract, and in most cases, a key illustration selected from the patent or patent application

NASA SBIR abstracts of 1991 phase 1 projects

The objectives of 301 projects placed under contract by the Small Business Innovation Research (SBIR) program of the National Aeronautics and Space Administration (NASA) are described. These projects were selected competitively from among proposals submitted to NASA in response to the 1991 SBIR Program Solicitation. The basic document consists of edited, non-proprietary abstracts of the winning proposals submitted by small businesses. The abstracts are presented under the 15 technical topics within which Phase 1 proposals were solicited. Each project was assigned a sequential identifying number from 001 to 301, in order of its appearance in the body of the report. Appendixes to provide additional information about the SBIR program and permit cross-reference of the 1991 Phase 1 projects by company name, location by state, principal investigator, NASA Field Center responsible for management of each project, and NASA contract number are included

The NASA SBIR product catalog

The purpose of this catalog is to assist small business firms in making the community aware of products emerging from their efforts in the Small Business Innovation Research (SBIR) program. It contains descriptions of some products that have advanced into Phase 3 and others that are identified as prospective products. Both lists of products in this catalog are based on information supplied by NASA SBIR contractors in responding to an invitation to be represented in this document. Generally, all products suggested by the small firms were included in order to meet the goals of information exchange for SBIR results. Of the 444 SBIR contractors NASA queried, 137 provided information on 219 products. The catalog presents the product information in the technology areas listed in the table of contents. Within each area, the products are listed in alphabetical order by product name and are given identifying numbers. Also included is an alphabetical listing of the companies that have products described. This listing cross-references the product list and provides information on the business activity of each firm. In addition, there are three indexes: one a list of firms by states, one that lists the products according to NASA Centers that managed the SBIR projects, and one that lists the products by the relevant Technical Topics utilized in NASA's annual program solicitation under which each SBIR project was selected

Lightning Imaging Sensor (LIS) for the Earth Observing System

Not only are scientific objectives and instrument characteristics given of a calibrated optical LIS for the EOS but also for the Tropical Rainfall Measuring Mission (TRMM) which was designed to acquire and study the distribution and variability of total lightning on a global basis. The LIS can be traced to a lightning mapper sensor planned for flight on the GOES meteorological satellites. The LIS consists of a staring imager optimized to detect and locate lightning. The LIS will detect and locate lightning with storm scale resolution (i.e., 5 to 10 km) over a large region of the Earth's surface along the orbital track of the satellite, mark the time of occurrence of the lightning, and measure the radiant energy. The LIS will have a nearly uniform 90 pct. detection efficiency within the area viewed by the sensor, and will detect intracloud and cloud-to-ground discharges during day and night conditions. Also, the LIS will monitor individual storms and storm systems long enough to obtain a measure of the lightning flashing rate when they are within the field of view of the LIS. The LIS attributes include low cost, low weight and power, low data rate, and important science. The LIS will study the hydrological cycle, general circulation and sea surface temperature variations, along with examinations of the electrical coupling of thunderstorms with the ionosphere and magnetosphere, and observations and modeling of the global electric circuit