A smartphone readout system for gold nanoparticle-based lateral flow assays: application to monitoring of digoxigenin (original) (raw)
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A New, High-Throughput HPLC/MS Assay for Therapeutic Level Monitoring of Digoxin in Human Plasma
A new high-throughput liquid chromatographic - mass spectrometric (LC-MS) assay for the quantification of digoxin in human plasma was developed and validated. The separation was performed on a Zorbax SB-C18 column under isocratic conditions using a 55:45 (v/v) mixture of methanol and 0.1% (v/v) formic acid in 10 μM sodium acetate at 45 ºC with a flow rate of 1 mL/min. The detection of digoxine was performed in SIM mode (m/z 803.5). The human plasma samples (0.2 mL) were deproteinized with 7% perchloric acid in water and aliquots of 20 μL from supernatants obtained after centrifugation were directly injected into the chromatographic system. The method shows a good linearity (r > 0.9926), precision (CV > 18.9 %) and accuracy (bias > 5.6 %) over the range of 0.5-50 ng/mL plasma. The lower limit of quantification (LLOQ) was 0.5 ng/mL and the recovery was between 92-106 %. The method is not expensive, it needs a minimum time for plasma sample preparation and has a run-time of 2.3 min for instrument analysis (retention time of digoxin was 1.9 min). The developed and validated high-throughput method is very simple, rapid and efficient, with wide applications in clinical level monitoring, pharmacokinetics and bioequivalence studies.
Scientific Reports, 2024
Rapid and label-free detection of very low concentrations of biomarkers in disease diagnosis or therapeutic drug monitoring is essential to prevent disease progression in Point of Care Testing. For this purpose, we propose a multipurpose optical Bio-Micro-Electro-Mechanical-System (BioMEMS) sensing platform which can precisely measure very small changes of biomolecules concentrations in plasma-like buffer samples. This is realized by the development of an interferometric detection method on highly sensitive MEMS transducers (cantilevers). This approach facilitates the precise analysis of the obtained results to determine the analyte type and its concentrations. Furthermore, the proposed multipurpose platform can be used for a wide range of biological assessments in various concentration levels by the use of an appropriate bioreceptor and the control of its coating density on the cantilever surface. In this study, the present system is prepared for the identification of digoxin medication in its therapeutic window for therapeutic drug monitoring as a case study. The experimental results represent the repeatability and stability of the proposed device as well as its capability to detect the analytes in less than eight minutes for all samples. In addition, according to the experiments carried out for very low concentrations of digoxin in plasma-like buffer, the detection limit of LOD = 300 fM and the maximum sensitivity of S = 5.5 × 10 12 AU/M are achieved for the implemented biosensor. The present ultrasensitive multipurpose BioMEMS sensor can be a fullyintegrated, cost-effective device to precisely analyze various biomarker concentrations for various biomedical applications. Nowadays, chronic diseases or noncommunicable diseases are one of the most important cause of death in the world (74% of all deaths [WHO, 2019]). Furthermore, the treatment or control of these diseases are very expensive compared to the early diagnosis and prevention processes 1,2. Consequently, the development of highperformance and cost-effective transducers for label-free detection of specific biomarkers, particularly for chronic diseases, is incredibly appealing to improve the human life quality by controlling the disease progression and drug monitoring for dosage adjustment in point-of-care (POC) testing. This may also decrease the total health cost expenditures of the worldwide health systems 3-5. In this context, biosensing technologies are good alternatives for time-consuming and expensive conventional methods of early diagnosis or therapeutic drug monitoring,
Analytical and Bioanalytical Chemistry, 2009
This work reports a comparative study of two automated flow-through fluorosensors for the determination of digoxin in serum samples: an immunosensor with an anti-digoxin polyclonal antibody as the reactive phase permanently immobilised on controlled-pore glass and a sensor with a selective reaction system based on a methacrylic molecularly imprinted polymer (MIP) synthesised by bulk polymerisation. The variables affecting the sensitivity and dynamic range of the sensors (e.g. the carrier and elution solutions, flow rates, pH and reagent concentrations) were optimized, and the binding characteristics of their reactive phases were compared in a competitive fluorescent assay. Digoxin was reproducibly determined by both sensors at the milligram per litre level (detection limit=1.20×10 −3 mg L −1 and RSD=4-7% for the immunosensor; detection limit= 1.7×10 −5 mg L −1 and RSD=1-2% for the MIP sensor). No cross-reactivity with digoxin-related compounds was seen for either sensor at a digoxin/interferent ratio of 1:100. The lifetime of the immunosensor was about 50 immunoassays; its shelf life, when unused, is about 3 months. The lifetime of the MIP sensor was over 18 months. Both sensors were used to determine the digoxin concentration of human serum samples with satisfactory results.
Design of Gold Nanoparticle Vertical Flow Assays for Point-of-Care Testing
Diagnostics
Vertical flow assays (VFAs) or flow-through assays have emerged as an alternate type of paper-based assay due to their faster detection time, larger sample volume capacity, and significantly higher multiplexing capabilities. They have been successfully employed to detect several different targets (polysaccharides, protein, and nucleic acids), although in a limited number of samples (serum, whole blood, plasma) compared to the more commonly known lateral flow assays (LFAs). The operation of a VFA relies mainly on gravity, coupled with capillary action or external force to help the sample flow through layers of stacked pads. With recent developments in this field, multiple layers of pads and signal readers have been optimized for more user-friendly operation, and VFAs have achieved a lower limit of detection for various analytes than the gold-standard methods. Thus, compared to the more widely used LFA, the VFA demonstrates certain advantages and is becoming an increasingly popular pl...
Therapeutic drug monitoring of digoxin–20 years of experience
Pharmacological Reports, 2017
Background: Digoxin is the oldest drug used in the pharmacotherapy of heart failure (HF). However, digoxin remains an important therapeutic option for patients with persistent symptoms of HF occurring despite the implementation of standard pharmacotherapy. Digoxin concentration serum (SCD) should equal 1-2 ng/ml. The aim of our study was to measure of SCD among the hospitalized patients as well as to determine the selected factors influencing the concentration of the digoxin in the blood. Methods: The presented research was based on a retrospective analysis including 2149 patients treated with digoxin and hospitalized between 1980 and 2000. Was used for the determination of SCD automatic analyzer TDX ABBOTT GmbH-fluorescence polarization immunoassay (FPIA), with therapeutic range for digoxin of 0.8-2.0 ng/ml. Results: Average SCD result in the study population was located within the therapeutic range and amounted 1.06 ng/ml (55.7 % of patients). Statistically significant differences in digoxin level were observed depending on the way of medicine administration (p=0.000001) and the daily amount (p=0.001). Moreover, statistically significant differences in digoxin level were observed depending on sex (p=0.00002). Conclusions: An elevated level of digoxin was observed in the case of patients who received the medication both orally and intravenously, together with an increase in the daily amount of digoxin doses. It was confirmed that an elevated digoxin level occurs in the course of treatment in the case of women.
Advantages of Highly Spherical Gold Nanoparticles as Labels for Lateral Flow Immunoassay
Sensors, 2020
The use of lateral flow immunoassays (LFIAs) for rapid on-site testing is restricted by their relatively high limit of detection (LoD). One possible way to decrease the LoD is to optimize nanoparticle properties that are used as labels. We compare two types of Au nanoparticles: usual quasispherical gold nanoparticles (C-GNPs), obtained by the Turkevich–Frens method, and superspherical gold nanoparticles (S-GNPs), obtained by a progressive overgrowth technique. Average diameters were 18.6–47.5 nm for C-GNPs and 20.2–90.4 nm for S-GNPs. Cardiomarker troponin I was considered as the target analyte. Adsorption and covalent conjugation with antibodies were tested for both GNP types. For C-GNPs, the minimal LoD was obtained with 33.7 nm nanoparticles, reaching 12.7 ng/mL for covalent immobilization and 9.9 ng/mL for adsorption. The average diameter of S-GNPs varied from 20.2 to 64.5 nm, which resulted in a decrease in LoD for an LFIA of troponin I from 3.4 to 1.2 ng/mL for covalent immobi...
Comparison of Single- and Mixed-Sized Gold Nanoparticles on Lateral Flow Assay for Albumin Detection
Biosensors
The sensitivity and reproducibility of the lateral flow assay can be influenced by multiple factors, such as the size of gold nanoparticles (GNPs) employed. Here, we evaluated the analytical performance of single-sized and mixed-sized GNPs using a simple lateral flow assay (LFA) platform. This platform was used as a model assay to diagnose albumin levels and demonstrate the analytical performance of single-sized and mixed-sized GNPs in LFA tests. Two sizes of GNPs@anti-bovine serum albumin (BSA) conjugate proteins were mixed at different ratios. The unique optical properties of the GNPs induced a distinguishing color-shedding effect on the single- and mixed-sized GNPs@anti-BSA conjugates interacting with the target analyte BSA spotted on the test line. The use of mixed-sized GNPs@anti-BSA conjugates enhanced signal relative to the 20 nm GNPs, and provided superior stability compared with solely employing the large GNPs (50 nm). The proposed platform in this study could provide an ef...
Lateral Flow Immunoassays - from Paper Strip to Smartphone Technology
Electroanalysis, 2015
Lateral flow immunoassays provide low cost, fast, portable and simple to use devices, with yes/no answers seen by the naked eye, that has found applications in agriculture, biomedicine, the environment, and food industries. Making these quantitative, via electrochemical or optical approaches, is the present day challenge, with a vision that one day, these will be connected to smart-phone technologies, which exhibit a promising digital platform for point-of-care diagnostics, mobile healthcare and bioanalytical needs. These devices are fully automated and equipped with a high resolution camera, a powerful processor with high storage capacity, wireless connectivity, real-time geo-tagging, secure data management, and cloud computing. CREATE program "Nanomaterials for Water and Energy Management" through MSE at the Nanyang Technological University. He has published in viral immunosensors (Ebola virus, West Nile virus, Rift Valley fever, Dengue, Hepatitis C) and has extensive experience in biosensors including chemiluminescent-based optical immunosensors to pathogen-elicited antibodies as well as amperometric immunosensors. He has developed new sensor configurations, such as establishing diagnostics based on luminescence emitted by primed neutrophils. His work also encompasses environmental toxicology such as monitoring water pollution via fiber-optic probes glowing in the presence of toxicants through their associated luminescent bacteria, or developing a nanometer particle-sensitive bioassay. His group has also participated in developing enzyme nanolithography, as well as ITO-based biochip configurations.
Biosensors and Bioelectronics, 2008
This work describes the development of a competitive flow-through FIA assay for digoxin using a molecularly imprinted polymer (MIP) as the recognition phase. In previous work, a number of non-covalent imprinted polymers were synthesised by "bulk" polymerisation. The digoxin binding and elution characteristics of these MIPs were then evaluated to obtain a highly selective material for integration into a sensor. The optimum MIP was synthesised by photo-initiated polymerisation of a mixture containing digoxin, MAA, EDGMA and AIBN in acetonitrile. The bulk polymer was ground and sieved and the template removed by Soxhlet extraction in MeOH/ACN. The MIP was packed into a flow cell and placed in a spectrofluorimeter to integrate the reaction and detection systems. The physical and chemical variables involved in digoxin determination by the sensor (nature and concentration of solution, flow rates, etc.) were optimised. Binding with the non-imprinted polymer (NIP) was also analysed. The new fluorosensor showed high selectivity and sensitivity, a detection limit of 1.7 × 10 −2 g l −1 , and high reproducibility (R.S.D. of 1.03% and 1.77% for concentrations of 1.0 × 10 −3 and 4.0 × 10 −3 mg l −1 , respectively). Selectivity was tested by determining the cross-reactivity of several compounds with structures analogous to digoxin. Under the assay conditions used, in which the potential interfering compounds were in concentrations 100 times higher than that of the analyte, no interference was recorded. The proposed fluorosensor was successfully used to determine digoxin concentration of human serum samples.
Journal of Immunological Methods, 2012
There is an increasing demand for convenient and accurate point-of-care tools that can detect and diagnose different stages of a disease in remote or impoverished settings. In recent years, lateral flow immunoassays (LFIA) have been indicated as a suitable medical diagnostic tool for these environments because they require little or no sample preparation, provide rapid and reliable results with no electronic components and thus can be manufactured at low costs and operated by unskilled personnel. However, even though they have been successfully applied to acute and chronic disease detection, LFIA based on gold nanoparticles, the standard marker, show serious limitations when high sensitivity is needed, such as early stage disease detection. Moreover, based on the lack of comparative information for label performance, significant optimization of the systems that are currently in use might be possible. To this end, in the presented work, we compare the detection limit between the four most used labels: colloidal-gold, silver enhanced gold, blue latex bead and carbon black nanoparticles. Preliminary results were obtained by using the biotin-streptavidin coupling as a model system and showed that carbon black had a remarkably low detection limit of 0.01 μg/mL in comparison to 0.1 μg/mL, 1 μg/mL and 1 mg/mL for silvercoated gold nanoparticles, gold nanoparticles and polystyrene beads, respectively. Therefore, as a proof of concept, carbon black was used in a detection system for Dengue fever. This was achieved by immobilizing monoclonal antibodies for the nonstructural glycoprotein (NS1) of the Dengue virus to carbon black. We found that the colorimetric detection limit of 57 ng/mL for carbon black was ten times lower than the 575 ng/mL observed for standard gold nanoparticles; which makes it sensitive enough to diagnose a patient on the first days of infection. We therefore conclude that, careful screening of detection labels should be performed as a necessary step during LFIA development in order to enhance the detection limit in a final test system.