Development towards a point-of-care system to monitor pregnancy and fertility biomarkers

The aim of this thesis was to develop a point of care (POC) device that could monitor progesterone and oestriol in saliva. These hormones have a key role to play in both female fertility and pregnancy. Understanding the concentration of progesterone in the body is key to understanding a patient’s fertility and pregnancy status. Combining progesterone and oestriol detection can give valuable insight into when labour may commence during pregnancy. This can be achieved by measuring the progesterone to oestriol ratio in saliva samples, throughout the pregnancy progesterone is the more dominant hormone but a few weeks before labour oestriol becomes the more dominant hormone. Saliva was chosen as the biological sample due to the ease of collection as well as it providing a better chemical understanding of the active hormonal concentrations compared to blood. Typical levels in saliva are 100 ng mL⁻¹ for progesterone and 0 - 5 ng mL⁻¹ for oestriol.The work presented began with the design of a chemiluminescence immunoassay which could be translated onto a microfluidic device, this approach would provide both good sensitivity and selectivity. Chemiluminescence was chosen as a detection system due to the high sensitivities that can be achieved with simple instrumentation where a CCD camera could be used to also obtain spatial information. The CL assay involved the chemical immobilisation of the antibodies onto glass slides. Silanisation with (3-aminopropyl)triethoxysilane (APTES) in combination with a N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide/N-hydroxysulfosuccinimide (EDC/sulfo-NHS) linker proved the most successful immobilisation method, with a LOD of 33 ±3 pg mL⁻¹ being achieved for progesterone in 10 mM phosphate buffered saline (PBS). This method however lacked reproducibility and did not transfer well on to polymer substrates or the microfluidic devices due to problems with the antibody immobilisation procedure. Immobilisation of anti-progesterone was then investigated on a range of electrode surfaces (Au, glassy carbon and ITO). This immobilisation procedure involved electrochemically depositing nitrobenzene onto the electrode surface followed by an electrochemical reduction of the nitro groups to the corresponding amine. To allow electrochemical detection ferrocene was tagged to the anti-progesterone antibodies to give a redox tag. The antibodies were then immobilised through an EDC/sulfo-NHS linkage. This method proved to be successful and very reproducible. By tagging ferrocene onto the antibody a rigid structure was achieved during the immobilisation procedure allowing the ideal antibody orientation, this process also allowed quantification of the concentration of antibodies on the surface (4.46 x10⁻⁷ mol m⁻²). Electrochemical based immunoassays were successfully carried with a 15 min incubation time for progesterone giving LODs of 1 pg mL⁻¹ for the gold and glassy carbon and 0.1 pg mL⁻¹ for the ITO. The ITO performed better than the other materials due to the electrode being uniformly flat enabling more efficient surface modifications. The methodology was also translated for use with artificial saliva with a LOD of 1.7 pg mL⁻¹ for progesterone.Once the electrochemical immobilisation platform had been shown to be successful this was taken forward as a potential route to carry out a CL immunoassay. This novel approach utilised the oxidised ferrocene tag on the antibody as the catalyst for the luminol CL reaction. A static system was devised in which the antibodies had been immobilised on to ITO using the electrochemical approach and the CL reagents added by pipette, LODs of 2.35 and 2.54 pg mL⁻¹ were obtained for progesterone and oestriol respectively in saliva after a 30 min incubation time. The static system could therefore be used as a POC device for these hormones meeting the aims of this thesis. The next step was then to start developing a more automated method within a flow cell. Initially the ITO electrode with the pre-prepared immobilised antibody and the oxidised ferrocene tag was incorporated into a macrofluidic device. The CL immunoassay was successfully carried out in the macrofluidic device although the LODs were an order of magnitude higher than those seen from the static system for both progesterone and oestriol in saliva due to the large volume of the flow cell. Finally the ITO electrode with pre-prepared immobilised antibody with the oxidised ferrocene tag was slotted into the microfluidic device that had been designed and measurement were made. There was however problems with the design and possible new designs are discussed

Laboratory evaluation of the Alere q point-of-care system for early infant HIV diagnosis

Introduction Early infant diagnosis (EID) and prompt linkage to care are critical to minimise the high morbidity and mortality associated with infant HIV infection. Attrition in the "EID cascade" is common; however, point-of-care (POC) EID assays with same-day result could facilitate prompt linkage of HIV-infected infant to treatment. Despite a number of POC EID assays in development, few have been independently evaluated and data on new technologies are urgently needed to inform policy. METHODS: We compared Alere q 1/2 Detect POC system laboratory test characteristics with the local standard of care (SOC), Roche CAP/CTM HIV-1 qualitative PCR in an independent laboratory-based evaluation in Cape Town, South Africa. Routinely EID samples collected between November 2013 and September 2014 were each tested by both SOC and POC systems. Repeat testing was done to troubleshoot any discrepancy between POC and SOC results. RESULTS: Overall, 1098 children with a median age of 47 days (IQR, 42-117) were included. Birth PCR (age <7 days) comprised of 8% (n = 92) tests while 56% (n = 620) of children tested as part of routine EID (ages 6-14 weeks). In the overall direct comparison, Alere q Detect achieved sensitivity of 95.5% (95% CI, 91.7-97.9%) and a specificity of 99.8% (95% CI, 99.1-100%). Following repeat testing of discordant samples and exclusion of any inconclusive results, the POC assay sensitivity and specificity were 96.9% (95% CI 93.4-98.9%) and 100% (lower 95% CI 98%) respectively. Among birth PCR tests the POC assay had slightly lower sensitivity (93.3% vs 96.5% in routine EID) and higher assay error rate (10% vs 5% in samples of older children, p = 0.04). CONCLUSION: Our results indicate this POC assay performs well for EID in the laboratory. The high specificity and thus high positive predictive value would suggest a positive POC result may be adequate for immediate infant ART initiation. While POC testing for EID may have particular utility for birth testing at delivery facilities, the lower sensitivity and error rate requires further attention, as does field implementation of POC EID technologies in other clinical care settings

Enabling High Value Care with A Point of Care Solution: the Australian Experience

Adopting patient-centric technology solutions is considered a critical enabler to enhance superior, high value healthcare delivery. Evidence from the literature underscores the simultaneous benefits of such an approach for enhancing the quality of care, increasing value and reducing associated costs. This study contributes to the current void in the literature by providing data from an implementation of a patient-centric solution that serves to deliver and support value based-care. Specifically, the presented study highlights how a point of care system can deliver high value patient-centric care across a healthcare group in Australia. The results from this qualitative study show that the examined point of care system supports patient-centric care by facilitating a high level of patient engagement and supporting key safety and quality care outcomes, as well as building a cultural shift towards patient-centric care as part of standard practice. The study has far reaching implications for both theory and practice

Information Systems and Healthcare XXI: A Dynamic, Client-Centric, Point-Of-Care System for the Novice Nurse

Nurse clinicians need to make complex decisions on a continual basis, while delivering cost-effective treatments. The rapid proliferation of medical and nursing knowledge complicates the decision-making process, particularly for novice nurses. We describe a Clinical Decision Support System (CDSS) for the novice nurse that combines evidence-based nursing knowledge with specific patient information to create a real-time guide through the nursing diagnostic care process. The goal of the paper is to describe how an appropriately designed and evidence-based CDSS can aid the nursing practice. An off-the-shelf handheld computer is utilized to deliver clinical knowledge to the nurse, via wireless link to a central server and a data repository. In describing the software architecture of the system, particular emphasis is paid to the issue of appropriate design by discussing the steps taken to address system extensibility, performance, reliability, and security, which are important factors in the design of a CDSS

Reconfigurable multiplexed point of Care System for monitoring type 1 diabetes patients

At the point of care (POC), on-side clinical testing allows fast biomarkers determination even in resource-limited environments. Current POC systems rely on tests selective to a single analyte or complex multiplexed systems with important portability and performance limitations. Hence, there is a need for handheld POC devices enabling the detection of multiple analytes with accuracy and simplicity. Here we present a reconfigurable smartphone-interfaced electrochemical Lab-on-a-Chip (LoC)with two working electrodes for dual analyte determination enabling biomarkers' selection in situ and on-demand. Biomarkers selection was achieved by the use of electrodepositable alginate hydrogels. Alginate membranes containing either glucose oxidase (GOx)or lactate oxidase (LOx)were selectively electrodeposited on the surface of each working electrode in around 4 min, completing sample measurement in less than 1 min. Glucose and lactate determination was performed simultaneously and without cross-talk in buffer, fetal bovine serum (FBS)and whole blood samples, the latter being possible by the size-exclusion filtration capacity of the hydrogels. At optimal conditions, glucose and lactate were determined in a wide linear range (0–12 mM and 0–5 mM, respectively)and with high sensitivities (0.24 and 0.54 μA cm −2 mM −1 , respectively), which allowed monitoring of Type-1 diabetic patients with a simple dual analysis system. After the measurement, membranes were removed by disaggregation with the calcium-chelator phosphate buffer. At this point, new membranes could be electrodeposited, this time being selective to the same or another analyte. This conferred the system with on-demand biomarkers’ selection capacity. The versatility and flexibility of the current architecture is expected to impact in POC analysis in applications ranging from homecare to sanitary emergencies.Peer reviewe

BIRS Course: RNA Vaccine Manufacture and Assessment of Regulatory Documents for RNA Vaccines

This paper is in three segments: (A) Segment on Vaccine Manufacture; (B) Segment on Ready to Use (RTU) Fluid Path for Compounded Sterile Preparations, mRNA Vaccines, and Phage Therapy, (C) Segment on Competency Framework for Addressing Regulatory Review These segments can be used separately or in combination. Additionally, they can be presented in any order. The time devoted to each segment depends on the depth of the course coverage. These segments are interrelated and describe how to make vaccines, how to manufacture vaccines with a point-of-care system built from ready-to-use parts; and how to regulate vaccines. This is a timely review because of the importance of vaccines for the treatment of diseases. It is hoped that it will lead to new approaches to vaccine manufacture and regulation

Analytical validation of the new plasma calibrated Accu-Chek (R) Test Strips (Roche Diagnostics)

peer reviewedBackground: The Accu-Chek Inform glucose monitor is a point-of-care system for testing blood glucose. New test strips, calibrated to deliver glucose plasma-like values, were launched on the market in May 2005. The aim of our study was to perform analytical validation of these new strips. Methods: We compared the new plasma strips with whole blood strips; results for the plasma strips with plasma values obtained using a clinical analyzer and with whole blood values given by the glucose electrode of a blood gas analyzer; and the influence of the type of blood (capillary or venous) on the results obtained by the glucose monitor with the plasma calibrated strips. Results: Plasma strips give on average 7% higher results than the previous whole blood strips. However, the results given by the plasma strips on capillary whole blood, even if well correlated, are not completely comparable with those given by an analyzer for venous plasma. Nevertheless, these plasma strips and the glucose electrode of a blood gas analyzer give comparable results. Conclusions: Accu-Chek Inform plasma strips are a good method for monitoring of blood glucose values in patients with diabetes

Handheld point-of-care system for rapid detection of SARS-CoV-2 extracted RNA in under 20 min

The COVID-19 pandemic is a global health emergency characterized by the high rate of transmission and ongoing increase of cases globally. Rapid point-of-care (PoC) diagnostics to detect the causative virus, SARS-CoV-2, are urgently needed to identify and isolate patients, contain its spread and guide clinical management. In this work, we report the development of a rapid PoC diagnostic test (<20 min) based on reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) and semiconductor technology for the detection of SARS-CoV-2 from extracted RNA samples. The developed LAMP assay was tested on a real-time benchtop instrument (RT-qLAMP) showing a lower limit of detection of 10 RNA copies per reaction. It was validated against extracted RNA from 183 clinical samples including 127 positive samples (screened by the CDC RT-qPCR assay). Results showed 91% sensitivity and 100% specificity when compared to RT-qPCR and average positive detection times of 15.45 ± 4.43 min. For validating the incorporation of the RT-LAMP assay onto our PoC platform (RT-eLAMP), a subset of samples was tested (n = 52), showing average detection times of 12.68 ± 2.56 min for positive samples (n = 34), demonstrating a comparable performance to a benchtop commercial instrument. Paired with a smartphone for results visualization and geolocalization, this portable diagnostic platform with secure cloud connectivity will enable real-time case identification and epidemiological surveillance

Investigation of Key Factors Affecting Quality of Patient Data from National Antiretroviral Therapy Electronic Medical Record System in Malawi

The Ministry of Health in Malawi implemented a National Antiretroviral Therapy Electronic Medical Record system currently deployed in over 150 health facilities. It thus expected quality and timely quarterly cohort reports. However, the raw electronic reports are rarely complete, accurate and consistent requiring cleaning hence being delayed. Such reports are now very critical under the COVID-19&nbsp; pandemic. Adopting a mixed-method approach, this study assessed the key factors that affect quality of data entered in the electronic medical records system and the reports produced by the system. The study interviewed 134 health-care workers in 17 sites and 10 Baobab Health Trust officers. Observations were conducted and secondary data analysed. The analysis shows that the EMRs lacks proper documentation and validation rules, making it hard to maintain and increasing chances of duplicate entry, respectively. Coupled with lack of trained personnel, it was revealed that one set of login credentials is used by multiple users and vital data elements being null compromising security and completeness, respectively. The electronic medical records system was not used at 40% of the sites as a point of care system hence being used as a back-data entry tool. Thus, there is need to revise the system to include necessary validations, security features, back data-entry form and data quality dashboards. Keywords: Electronic Medical Records system, Data Quality, System Quality, Information Qualit