Browsing by Author "Hemben, Aver"
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Item Open Access Development of immunosensors for the detection of malaria.(2017-05) Hemben, Aver; Tothill, Ibtisam E.Malaria is a disease of global importance caused by an Apicomplexan Plasmodium parasite and transmitted by adult female Anopheles mosquitoes. Malaria affects approximately 50% of the world’s population causing millions of deaths every year. Mostly affected are pregnant women and children under 5 years of age. Morbidity and mortality rates are on the decline in some areas. Despite control efforts the disease continues to affect productivity. Productivity can be increased by early detection. Methods for malaria detection include blood film microscopy, immunochromatographic, serological and molecular tests. Blood film microscopy shows the highest sensitivity and specificity when used by trained personnel with reliable instruments. It is however time-consuming and cannot be applied as a point-of-care diagnostic method. Two electrochemical immunosensors for malaria biomarkers Plasmodium falciparum histidine rich protein 2 (PfHRP 2) and parasite L-Lactate dehydrogenase (LDH) were developed in this work for the detection and quantification of Plasmodium species. The methods were based on screen-printed gold electrodes (SPGEs) with on board carbon counter and silver /silver chloride (Ag / AgCl) pseudo-reference electrode. The first stage of the work involved comparison by characterization of the bare SPGEs using potassium ferricyanide. Electrochemical techniques were used to compare bare and self-assembled monolayers of mercaptoundecanoic acid (MUA) and 3,3´- dithiodipropionic acid (DTDPA) against bare SPGE. The optimal sensor was then used for antibody attachment. For the second stage of the work, adsorption was investigated for capture antibody immobilization on the SPGE. HuCAL monoclonal antibodies against PfHRP 2 conjugated to the electroactive enzyme horseradish peroxidase (HRP) were then applied for signal generation. Electrochemical measurements were conducted using 3,3´ 5,5´-tetramethylbenzidine dihydrochloride and hydrogen,peroxide (TMB / H₂O₂) as the mediator / substrate system at potential of -0.2 V. The sensors utilized sandwich enzyme-linked immunosorbent assay (ELISA),format with HuCAL monoclonal antibodies against Plasmodium immobilized on the gold working electrode. The developed biosensor was capable of detecting sub-microscopic Plasmodium infection with a linear range from 1 to 100 ng mL⁻¹ and a limit of detection (LOD) as low as 2.14 ng mL⁻¹ and 2.95 ng mL⁻¹ for PfHRP 2 in buffer and serum assays respectively. When compared with AuNP enhanced assays, the LOD was 36 pg mL⁻¹ and 40 pg mL⁻¹.. Another biomarker Plasmodium falciparum parasite Lactate dehydrogenase (LDH) was also investigated and another sensor developed using a sandwich assay similar to the PfHRP 2 sensor, but incorporating different antibodies against LDH. LOD 1.80 ng mL⁻¹ and 0.70 ng mL⁻¹ for LDH was obtained in buffer and serum assays. When compared with AuNP enhanced assays, the LOD was 19 pg mL⁻¹ and 23 pg mL⁻¹ respectively. As part of the work, culture medium supernatant containing PfHRP 2 and LDH was used to compare the immunosensor sensitivity for the pan-malaria antigen LDH. Sensitivity of the immunosensor was compared against commercially available Plasmodium immunochromatographic (ICT) kits: OptiMAL-IT and BinaxNOW Malaria kits. The optimized immuno-electrochemical biosensor detected the antigen at 0.002 % parasitaemia whereas the OptiMAL-IT ICT was only able to detect the LDH antigen when concentrations were of 2% parasitaemia. BinaxNOW ICT detected both the LDH and PfHRP 2 antigens in concentrations of 4% parasitaemia and showed negative reading at 0.5%parasitaemia in both synchronized and asynchronized samples. This study has developed two highly sensitive, portable and low cost malaria immunosensors for the first time on JD SPGEs. LDH immunosensor detects all Plasmodium species while PfHRP 2 immunosensor is specific for the detection of Plasmodium falciparum biomarker. Both immunosensors detect quantifiable, sub-microscopic levels of the biomarkers with sensitivities higher than the ICT tests. The immunosensors are therefore recommended for field trial.Item Open Access Metallic nano carrier complex targeting neuroendocrine prostate cancer(Cranfield University, 2019-05-09 10:33) Hemben, Aver; Chianella, Iva; Leighton, GlennPoster presented at Cranfield University’s 2019 Manufacturing Doctoral Community event.Item Open Access Metallic nanocarrier complex targeting neuroendocrine prostate cancel cells.(2022-09) Hemben, Aver; Chianella, Iva; Leighton, GlennIron oxide nanoparticles were successfully produced by inert gas condensation using the Mantis NanoGen Trio system. The nanoparticles were produced via a novel approach using the Physical Vapour Deposition (PVD) method to condense iron oxide nanoparticles of small size and small size distribution of 1-6 nm diameter. The sputtered nanoparticles were soft-landed on a polyethylene glycol-coated silicon wafer, then dispersed in RNAse free water (PEG-IONPs) to functionalise them for biocompatibility and use in nanocarrier synthesis. The sputtered iron oxide nanoparticles were characterised by transmission electron microscopy, atomic force microscopy, nanoparticle tracking analysis, dynamic light scattering and magnetic resonance imaging. The different techniques demonstrated that whereas the IONPs themselves were less than 10 nm (seen by TEM) the PEG-IONPs demonstrated a size range slightly different according to the technique used for its evaluation (e.g.47 ± 22.75 nm by DLS and 143 ± 100 nm by NTA) with the variation most likely due to the high concentration of PEG present in solution. In parallel to sputtering IONPs, commercial iron oxide nanoparticles (CIONPs) were used to demonstrate and optimise the attachment of a therapeutic drug (siRNA) to the nanoparticles in 1:1 polyethylene glycol and polyethyleneimine. Then the release profile of the attached siRNA from CIONPs in PBS buffer pH 6.4 and 7.4 was studied. While using a sample of PBS/PEG/PEI, approximately 75% of siRNA was attached to CIONPs and approximately 86% siRNA was released in PBS after 300 minutes of incubation at 37°C. The research findings demonstrate the potential use of the synthesised nanoparticles and nanocarrier complex in targeted drug deliver to neuroendocrine prostate cells.