Development and evaluation of point-of-care diagnostic technologies for providers and consumers

Point-of-care (POC) diagnostic technologies aim to expand access to traditional laboratory-based testing to near-patient settings. These settings can range from emergency or intensive care-units (ICUs) in the United States, to remote health posts in sub Saharan Africa. Differences in budget and infrastructure play a role in characterizing the wide array of possible “near patient” settings and must be taken into consideration in the engineering design process. In this dissertation we use translational engineering to develop practical and accessible microfluidic POC immunoassays for diverse settings, that include both provider and consumer facing applications. First, we examined Lyme Disease in the U.S., where existing diagnostic technologies face the challenge of rapid and accurate serodiagnosis in the face of largely non-specific clinical symptoms. We developed a multiplexed rapid test that could replicate enzyme-linked immunosorbent assay (ELISA) performance for Lyme Disease diagnosis. After screening candidate biomarkers, we evaluated performance of the multiplexed microfluidic test against ELISA using clinical serum samples and illustrated the potential to streamline current clinical algorithms requiring two immunoassays (ELISA and Western Blot) into one standalone test suitable for physician’s offices or urgent care clinics in the U.S. We also showed exploratory work towards a similar multiplexed test design for another bacterial spirochete infection, Leptospirosis. Next, we built on previous work towards a POC HIV-syphilis antenatal screening tool, to develop a smartphone-integrated, microfluidic assay for healthcare workers to use in low resource settings. The low-cost ($34), re-usable device (“smartphone dongle”) costs$34 to produce and provides results in 15 minutes. In this work, we focus on assay development efforts undertaken towards development of a fully integrated POC product suitable for deployment in the field, with practical considerations for the use of fingerstick blood, stability, scale-up and transport. We also streamlined the number of manual steps for end-user operation, through the use of lyophilized secondary antibodies, preloaded reagents on cassette, and an automatic result readout. While laboratory demonstration with clinical samples is important for initial characterization of POC devices, field evaluation reveals diagnostic performance under real-world conditions. We tested the device in the hands of minimally trained healthcare workers in Rwanda and saw comparable performance to other immunoassays run under field conditions. We also performed a follow-up pilot field study in Rwanda to evaluate the feasibility of the smartphone dongle platform for self-testing by patients/consumers in a low-resource setting, one of the most challenging use-cases for POC devices. Finally, we sought to integrate intellectual frameworks from behavioral research and user-experience (UX) design in creating a new framework for evaluation of consumer-facing microfluidic devices, specifically towards HIV home-testing in the U.S. While overall rates of HIV are decreasing in the U.S., the population of gay, bisexual and other men who have sex with men (MSM) are disproportionately affected. Self-testing products for sexually transmitted infection (STI) testing could address unmet needs for these target populations in both increasing access and frequency of testing, as well as integrating use with sexual partners for early diagnosis or even prevention. We worked with a cohort of MSMs at high risk for HIV/STI transmission in New York City, and performed for the first time, a structured assessment of completely naïve users interacting with a smartphone interfaced microfluidic diagnostic device (“SMARTtest”). We integrated UX design value model of device usability, credibility, accessibility and acceptability into our evaluation framework, which influence user’s information, knowledge, motivation and behavioral skills towards engaging with a prevention method (“IMB” model). Thus far, such frameworks have rarely been applied to other consumer health monitoring devices, including microfluidic POC devices. As the microfluidic field moves towards more field demonstrations of devices, more untrained and minimally trained users will have access to such tools. It is important to understand how they use devices, what the device failure points are and what the most relevant design features are to spur user adoption and meaningful usage. Underlying our work in creating accessible and practical POC immunoassay tools for infectious disease detection, is the illustration of the translational development roadmap from proof-of-concept assay development to field studies and user-based evaluations for intended end-use settings that range from U.S. based primary care clinics, rural health centers in low-resource settings as well as self-testing environments in both. Incorporating an understanding of the target use-case setting is critical in translating technologies for clinical use, whether in the infrastructure and services that are available, or end-user needs and constraints such as clinical workflow patterns, level of technical expertise and perceptions of usefulness and value. We show how user/use-case focused application of downstream translational engineering and testing informs upstream design choices and accelerates development of POC devices for real-world use. The sum of this work aims to illustrate tenets of translational engineering design and testing to advance insight into building POC products that are poised for greater adoption by target end users, whether they are health providers or consumers

Phlebot: The Robotic Phlebotomist

Phlebotomy is a routine task, performed over a billion times annually in the United States alone, that is essential for proper diagnosis and treatment. We designed and constructed Phlebot, a robotic assistive device that uses near- infrared imaging and force-feedback to guide a needle into a forearm vein for blood sample collection or intravenous catheterization. Through initial validation on phantoms, we show that it is feasible to automate phlebotomy reliably. We envision the device to be a first major step towards more affordable point-of-care testing and diagnostic healthcare systems. In the long term, we expect that Phlebot will expedite healthcare delivery and drastically reduce needle stick injuries, instances of hemolysis, and infections caused by blood-borne pathogens

Expanding accessibility of diagnostics through miniaturized technologies

There is a disproportionate burden of disease (measured in daily-adjusted life years, or DALYs) in low-income countries. Much of this disparity is due to infectious diseases: 53% of DALYs in Africa are due to infectious diseases, compared with only 3% in the American continents. This disparity is largely due to differences in electrical and transport infrastructure as well as access to skilled personnel and monetary resources. Current diagnostic solutions are primarily designed for high-resource settings and therefore these solutions cannot be easily translated to a lower-resource setting. In order to tackle this health disparity, new solutions must be designed specifically for a lower-resource setting. In this dissertation, we take a translational approach to engineering appropriate diagnostics for resource-limited settings. First, we develop a handheld smartphone accessory to perform an assay similar to enzyme-linked immunosorbent assay (ELISA), traditionally a laboratory-based test. In 15 minutes, it provides an objective diagnostic readout important for minimal training, while using an average of 1.6mW of power and costing only $34. We further develop the device to provide a quantitative hemoglobin measurement simultaneously with an HIV immunoassay, for use in antenatal care screening. The multiplexing two assay types that are clinically relevant has the potential to streamline workflow. While specifications can be demonstrated in the laboratory, the true test of the device must be performed in the field. We brought our smartphone accessory to three health centers in Kigali, Rwanda to be used by healthcare workers with no prior experience in ELISA. After a short 30 minute training, the healthcare workers were able to obtain diagnostic results comparable to other immunoassays run under field conditions. With a simple and user-friendly design, we sought to further expand the usage of our device as a self-testing device, having patients test themselves. Lastly, we explore manufacturable thermoplastics as a material for a microfluidic diagnostic for nucleic acid detection. The sum of this work aims to gain insight into methods of design, testing, and implementation of translational design

PhleAid™ : Automated Phlebotomy Assistant

PhleAid\u27s goal is to minimize pre-analytical errors and simplify the phlebotomy process. PhleAid implements a combination of three subsystems: a vial selection, handling, labeling and blood verification system. The selection system chooses the appropriate vial for each test through a graphical input and decision system that utilizes a microprocessor and database. The handling and labeling system minimizes user contact with vials by implementing our designed hardware and stepper motors. Additionally, a barcode scanner is implemented to identify each blood vial. Finally, the blood volume verification system utilizes photodiodes, LEDs and amplifiers to ensure sufficient the blood volume. Overall these subsystems work together to achieve our goal

Aerospace medicine and biology: A continuing bibliography with indexes (supplement 357)

This bibliography lists 186 reports, articles and other documents introduced into the NASA Scientific and Technical Information System during Dec. 1991. Subject coverage includes: aerospace medicine and physiology, life support systems and man/system technology, protective clothing, exobiology and extraterrestrial life, planetary biology, and flight crew behavior and performance

Research and technology: 1986 annual report of the Lyndon B. Johnson Space Center

Johnson Space Center accomplishments in new and advanced concepts during 1986 are highlighted. Included are research funded by the Office of Aeronautics and Space Technology; Solar System Exploration and Life Sciences research funded by the Office of Space Sciences and Applications; and Advanced Programs tasks funded by the Office of Space Flight. Summary sections describing the role of the Johnson Space Center in each program are followed by one-page descriptions of significant projects. Descriptions are suitable for external consumption, free of technical jargon, and illustrated to increase ease of comprehension

Field Immune Assessment during Simulated Planetary Exploration in the Canadian Arctic

Dysregulation of the immune system has been shown to occur during space flight, although the detailed nature of the phenomenon and the clinical risks for exploration class missions has yet to be established. In addition, the growing clinical significance of immune system evaluation combined with epidemic infectious disease rates in third world countries provides a strong rationale for the development of field-compatible clinical immunology techniques and equipment. In July 2002 NASA performed a comprehensive field immunology assessment on crewmembers participating in the Haughton-Mars Project (HMP) on Devon Island in the high Canadian Arctic. The purpose of the study was to evaluate mission-associated effects on the human immune system, as well as to evaluate techniques developed for processing immune samples in remote field locations. Ten HMP-2002 participants volunteered for the study. A field protocol was developed at NASA-JSC for performing sample collection, blood staining/processing for immunophenotype analysis, wholeblood mitogenic culture for functional assessments and cell-sample preservation on-location at Devon Island. Specific assays included peripheral leukocyte distribution; constitutively activated T cells, intracellular cytokine profiles and plasma EBV viral antibody levels. Study timepoints were L-30, midmission and R+60. The protocol developed for immune sample processing in remote field locations functioned properly. Samples were processed in the field location, and stabilized for subsequent analysis at the Johnson Space Center in Houston. The data indicated that some phenotype, immune function and stress hormone changes occurred in the HMP field participants that were largely distinct from pre-mission baseline and post-mission recovery data. These immune changes appear similar to those observed in Astronauts following spaceflight. The sample processing protocol developed for this study may have applications for immune assessment during exploration-class space missions or in remote terrestrial field locations. The data validate the use of the HMP as a ground-based spaceflight/planetary exploration analog for some aspects of human physiology

Towards early hemolysis detection: a smartphone based approach

Os especialistas em diagnóstico in vitro (IVDs) têm confiado maioritariamente na inspeção visual (ótica) manual e, em segundo lugar, em sensores óticos ou câmaras embutidas ou dispositivos médicos incorporados que suportam o exame da qualidade da amostra na fase pré-analítica. Com o aumento dos volumes de amostras para serem processadas e dos respetivos dados complexos gerados por esse processamento, aquelas técnicas tornaram-se cada vez mais difíceis de utilizar, ou os respetivos resultados não ficam imediatamente disponíveis. Para superar as complexidades impostas por tais técnicas tradicionais, o aumento do uso de dispositivos móveis e algoritmos de processamento de imagem no setor de saúde abriu caminho para a constituição de novos casos de uso baseados em análises móveis de amostras, pois fornecem uma interação simples e intuitiva com objetos gráficos familiares que são mostrados no ecrã dos smartphones. As interfaces gráficas e as técnicas de interação suportadas por dispositivos móveis podem pois proporcionar ao especialista em IVD uma série de vantagens e valor agregado devido à maior familiaridade com estes dispositivos e à grande acessibilidade que evidenciam atualmente, tendo o potencial de facilitar as análises de amostras. No entanto, o uso sistemático de dispositivos móveis no setor da saúde encontra-se ainda numa fase muito incipiente, em particular na área de IVD. Nesta tese, propõe-se conceber e discutir a arquitetura, a conceção e a implementação de um protótipo de uma aplicação móvel para smartphone (designada por "HemoDetect") que implementa um conjunto sugerido de algoritmos, interfaces e técnicas de interação que foram desenvolvidos com o objetivo de contribuir para a compreensão de técnicas mais eficientes para ajudar a detetar a hemólise, um processo que designa a rotura de glóbulos vermelhos (eritrócitos) e libertação do respetivo conteúdo (citoplasma) para o fluído circundante (por exemplo, plasma sanguíneo), complementando-as com estatísticas e medições de laboratório, mostrando a utilização de um protótipo durante experiências, permitindo assim chegar-se a um conceito viável que permita apoiar eficazmente a deteção precoce de hemólise.In Vitro Diagnostics (IVDs) specialists have been firstly relying on manual visual (optical) inspection and, secondly, on optical sensors or cameras embedded or built-in medical devices which support the examination of sample quality in pre-analytical phase. With increasing sample processing volumes and their generated complex data, these techniques have become increasingly difficult or results are not readily available. In order to overcome the complexities posed by these traditional techniques, the increased usage of mobile devices and algorithms in the healthcare industry paves the way into shaping new use cases and discovery of mobile analysis of samples, as they provide a user-friendly and familiar interaction with objects displayed on their screens. The interfaces and interaction techniques rendered by mobile devices, bring, to the IVD specialist, a number of advantages and added value due to increased familiarity with the devices or their accessibility, which is made easier. However, they are at the beginning of their journey in the healthcare industry, in particular in the IVD and point-of-care areas. In this thesis, the proposal is to discover and discuss the architecture, design and implementation of a smartphone prototype app (called “HemoDetect”) with its algorithms, interfaces and interaction techniques which was developed to help detect hemolysis which represents the rupture of red blood cells (erythrocytes) and release of their contents (cytoplasm) into surrounding fluid (e.g. blood plasma), and complementing it with from-the-lab statistics and measurements showing its utilization during experiments, which ultimately may be a feasible concept that could support early hemolysis detection.Les spécialistes du diagnostic in vitro (DIV) se sont d'abord appuyés sur l'inspection visuelle (optique) manuelle et, ensuite, sur des capteurs optiques ou des caméras intégrées ou intégrées à des dispositifs médicaux qui facilitent l'examen de la qualité des échantillons en phase pré-analytique. Avec l'augmentation des volumes de traitement des échantillons et des données complexes générées, ces techniques sont devenues de plus en plus difficiles ou les résultats ne sont pas facilement disponibles. Afin de surmonter les complexités posées par ces techniques traditionnelles, l'utilisation croissante des appareils mobiles et des algorithmes dans le secteur de la santé ouvre la voie à la définition de nouveaux cas d'utilisation et à la découverte d'analyses d'échantillons mobiles, car ils fournissent une interaction conviviale et familière. avec des objets affichés sur leurs écrans. Les interfaces et les techniques d'interaction rendues par les appareils mobiles apportent au spécialiste des dispositifs de DIV un certain nombre d'avantages et de valeur ajoutée en raison d'une familiarisation accrue avec les appareils ou de leur accessibilité, ce qui est facilité. Cependant, ils sont au début de leur parcours dans le secteur de la santé, en particulier dans le domains des DIV et point-of-care. Dans cette thèse, la proposition est de découvrir et de discuter de l’architecture, de la conception et de la mise en oeuvre d’une application pour smartphone (appelée «HemoDetect») avec ses algorithmes, interfaces et techniques d’interaction, qui a été développée pour aider à détecter l’hémolyse qui représente une rupture des globules rouges (érythrocytes) et la libération de leur contenu (cytoplasme) dans le liquide environnant (par exemple, le plasma sanguin), en le complétant par des statistiques de laboratoire et des mesures montrant son utilisation au cours des expériences, ce qui pourrait finalement être un concept réalisable qui pourrait permettre une détection précoce de l'hémolyse

Applications of Mass Spectrometry in Proteomics and Pharmacokinetics

Tremendous technology improvements of the last decades has given mass spectrometry a more and more expanding role in the study of a wide range of molecules: from the identification and quantification of small molecular weight molecules to the structural determination of biomacromolecules. Many are the fields of application for this technique and the various versions of it. In the present study three different applications have been explored. The first application is a pharmacokinetics study of anticancer drug Gemcitabine and its principal metabolite, where the role of the LC-MS/MS is essential both for the selectivity of the detection of the small analytes and the sensitivity enhanced by multi-reaction monitoring experiments. The design of the study involved the collection of several blood samples at selected times and from patients that would have met certain eligibility criteria. The ESI demonstrated to be the most suitable approach and it provided the necessary data to conclude that toxicity of Gemcitabine did not increase when administered at FDR (Fixed Dose Rate) infusion in patients with impaired hepatic function. The second application describes an example of how MS represents a powerful tool in cancer research, from serum profiling study with high resolution MALDITOF and bioinformatic analysis, to the identification of potential biomarker through peak identification. Almost 400 serum sample – homogeneously distributed between biopsy confirmed ovarian cancer and high risk serum samples – were analyzed on a high resolution MALDI-TOF instrument after automated reverse phase magnetic beads separation. The high throughput data have undergone sophisticated bioinformatic procedures that lead to a list of upand down-regulated peaks, although identification studies were possible only for those peaks that showed a good reproducibility. One down-regolated peak has been identified using the LC-MS/MS technique. The identified peak confirmed a basic role of fibrinogen in the ovarian cancer; the other four peaks that have been identified as down-regulated showed an absolutely not satisfactory ionization in electro-spray, therefore further analysis will be performed on these analytes in order to determinate their amino acidic sequence. The most suitable technique seems to be MALDI-TOF/TOF mass spectrometry, since the peptides already showed a good degree of ionization in MALDI. The third and last study belongs to a quite new field, which is the combination of immuno precipitation assays with MALDI-TOF (Immuno Precipitation Mass Spectrometry, IPMS) experiments in order to evaluate the specificity of a series of monoclonal antibodies to specific antigen. The automated assay that has been developed provides structural information about the antigen that binds the monoclonal antibody to be tested and previously conjugated to the surface of magnetic beads, ideal support for robotic automation. IPMS showed its potential as a complementary tool of crucial importance in the selection of the monoclonal antibody for the development of ELISA based assay to be applied in the screening of a consistent number of human specimens for the clinical validation of proteins indicated in literature as potential biomarkers. Mass spectrometry in association with fractionation techniques, such as liquid or magnetic beads chromatography, is a very flexible tool in the cancer research field. Further improvement in the instrumentation and in the technology will bring always more and more results to be confident in

Applications of Mass Spectrometry in Proteomics and Pharmacokinetics

Tremendous technology improvements of the last decades has given mass spectrometry a more and more expanding role in the study of a wide range of molecules: from the identification and quantification of small molecular weight molecules to the structural determination of biomacromolecules. Many are the fields of application for this technique and the various versions of it. In the present study three different applications have been explored. The first application is a pharmacokinetics study of anticancer drug Gemcitabine and its principal metabolite, where the role of the LC-MS/MS is essential both for the selectivity of the detection of the small analytes and the sensitivity enhanced by multi-reaction monitoring experiments. The design of the study involved the collection of several blood samples at selected times and from patients that would have met certain eligibility criteria. The ESI demonstrated to be the most suitable approach and it provided the necessary data to conclude that toxicity of Gemcitabine did not increase when administered at FDR (Fixed Dose Rate) infusion in patients with impaired hepatic function. The second application describes an example of how MS represents a powerful tool in cancer research, from serum profiling study with high resolution MALDITOF and bioinformatic analysis, to the identification of potential biomarker through peak identification. Almost 400 serum sample – homogeneously distributed between biopsy confirmed ovarian cancer and high risk serum samples – were analyzed on a high resolution MALDI-TOF instrument after automated reverse phase magnetic beads separation. The high throughput data have undergone sophisticated bioinformatic procedures that lead to a list of upand down-regulated peaks, although identification studies were possible only for those peaks that showed a good reproducibility. One down-regolated peak has been identified using the LC-MS/MS technique. The identified peak confirmed a basic role of fibrinogen in the ovarian cancer; the other four peaks that have been identified as down-regulated showed an absolutely not satisfactory ionization in electro-spray, therefore further analysis will be performed on these analytes in order to determinate their amino acidic sequence. The most suitable technique seems to be MALDI-TOF/TOF mass spectrometry, since the peptides already showed a good degree of ionization in MALDI. The third and last study belongs to a quite new field, which is the combination of immuno precipitation assays with MALDI-TOF (Immuno Precipitation Mass Spectrometry, IPMS) experiments in order to evaluate the specificity of a series of monoclonal antibodies to specific antigen. The automated assay that has been developed provides structural information about the antigen that binds the monoclonal antibody to be tested and previously conjugated to the surface of magnetic beads, ideal support for robotic automation. IPMS showed its potential as a complementary tool of crucial importance in the selection of the monoclonal antibody for the development of ELISA based assay to be applied in the screening of a consistent number of human specimens for the clinical validation of proteins indicated in literature as potential biomarkers. Mass spectrometry in association with fractionation techniques, such as liquid or magnetic beads chromatography, is a very flexible tool in the cancer research field. Further improvement in the instrumentation and in the technology will bring always more and more results to be confident in