A rapid magnetic bead-based immunoassay for sensitive determination of diclofenac.
Ontology highlight
ABSTRACT:
SUBMITTER: Ecke A
PROVIDER: S-EPMC8761716 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Immunological methods to detect SARS-CoV-2 seroconversion in humans are important to track COVID-19 cases and the humoral response to SARS-CoV-2 infections and immunization to future vaccines. The aim of this work was to develop a simple chromogenic magnetic bead-based immunoassay which allows rapid, inexpensive, and quantitative detection of human antibodies against SARS-CoV-2 in serum, plasma, or blood. Recombinant 6xHis-tagged SARS-CoV-2 Nucleocapsid protein was mobilized on the surface of Ni2+ magnetic beads and challenged with serum or blood samples obtained from controls or COVID-19 cases. The beads were washed, incubated with anti-human IgG-HPR conjugate, and immersed into a solution containing a chromogenic HPR substrate. Bead transfer and homogenization between solutions was aided by a simple low-cost device. The method was validated by two independent laboratories, and the performance to detect SARS-CoV-2 seroconversion in humans was in the same range as obtained using the gold standard immunoassays ELISA and Luminex, though requiring only a fraction of consumables, instrumentation, time to deliver results, and volume of sample. Furthermore, the results obtained with the method described can be visually interpreted without compromising accuracy as demonstrated by validation at a point-of-care unit. The magnetic bead immunoassay throughput can be customized on demand and is readily adapted to be used with any other 6xHis tagged protein or peptide as antigen to track other diseases.
| S-EPMC7860136 | biostudies-literature
Project description:Pharmacologically active compounds are often detected in wastewater and surface waters. The nonsteroidal anti-inflammatory drug diclofenac (DCF) was included in the European watch list of substances that requires its environmental monitoring in the member states. DCF may harmfully influence the ecosystem already at concentrations ≤ 1 μg L-1. The fast and easy quantification of DCF is becoming a subject of global importance. Fluorescence polarization immunoassay (FPIA) is a homogeneous mix-and-read method which does not require the immobilization of reagents. FPIA can be performed in one phase within 20-30 min, making it possible to analyse wastewater without any complicated pre-treatment. In this study, new tracer molecules with different structures, linking fluorophores to derivatives of the analyte, were synthesized, three homologous tracers based on DCF, two including a C6 spacer, and one heterologous tracer derived from 5-hydroxy-DCF. The tracer molecules were thoroughly assessed for performance. Regarding sensitivity of the FPIA, the lowest limit of detection reached was 2.0 μg L-1 with a working range up to 870 μg L-1. The method was validated for real wastewater samples against LC-MS/MS as reference method with good agreement of both methods. Graphical abstract.
| S-EPMC7813709 | biostudies-literature
Project description:Cell-surface proteins represent an important class of molecules for therapeutic targeting and defining cellular phenotypes. However, their enrichment and detection via mass spectrometry-based proteomics remains challenging due to low abundance, posttranslational modifications, hydrophobic regions, and processing requirements. To improve the identification of cell-surface proteins via their corresponding N-linked glycopeptides (N-glycopeptides), we optimized a Cell-Surface Capture (CSC) workflow which incorporates magnetic bead-based processing. Using this approach, we evaluated labeling conditions (biotin tags and catalysts), enrichment specificity (streptavidin beads), missed cleavages (lysis buffers), non-enzymatic deamidation (digestion and de-glycosylation buffers), and data acquisition methods. Our findings support the use of alkoxyamine-PEG4-biotin plus 5-methoxy-anthranilic acid and streptavidin magnetic beads for maximal N-glycopeptide detection. Furthermore, single-pot solid-phased-enhanced sample-preparation (SP3) circumvented the need to isolate cell membranes by affording the use of strong detergents and chaotropes for protein extraction. Notably, with semi-automated processing, sample handling was simplified and between ~600-900 N-glycoproteins were identified from only 25-200µg of HeLa protein. Overall, the improved efficiency of the magnetic-based CSC workflow allowed us to identify both previously reported and novel N-glycosites with less material and high reproducibility, and should help advance the field of surfaceomics by providing insight in cellular phenotypes not previously documented.
2024-01-21 | PXD043678 | Pride
Project description:Colorectal cancer (CRC) is the second leading cause of cancer deaths in the US and Western world. Despite increased screening and advances in treatment, the mortality rate (ca. 50,000/year) and high national health-care burden for CRC are likely to remain high unless an effective non-invasive screening test for CRC is instituted for a large segment of the population. Blood-based protein biomarkers hold great promise for early disease diagnosis and personalized medicine; yet robust and reproducible multiplexing platforms and methodologies have lagged behind their genomic counterparts. Here, we report the development of a novel, multiplexed, hybrid immunoassay for CRC that is formatted on barcoded VeraCode™ micro-beads, which have until now only been used for genomic assays. The method combines a sandwich immunoassay format for detection of serum protein biomarkers with an antigen assay for autoantibody detection. The serum protein biomarkers CEA and GDF15 as well as autoantibodies to the p53 tumor associated antigen (TAA) were used to exemplify the method. This multiplex biomarker panel was configured to run on Illumina's holographically barcoded VeraCode™ micro-bead platform, which is capable of measuring hundreds of analytes simultaneously in a single well from small volumes of blood (<50 ?L) using a 96-well industry standard microtiter plate. This novel use of the VeraCode™ micro-bead platform translates into a potentially low volume, high throughput, multiplexed assay for CRC, for the purposes of biomarker validation, as well as patient screening, diagnostics and prognostics. In an evaluation of a 186 patient sera training set (CRC and normal), we obtained a diagnostic sensitivity of 54% and a specificity of 98%. We anticipate that by expanding and refining the biomarkers in this initial panel, and performing more extensive clinical validations, such an assay could ultimately provide a basis for CRC population screening to complement the more invasive, expensive and low throughput (but highly sensitive and specific) colonoscopy.
| S-EPMC3867820 | biostudies-literature
Project description:In this study, a fluorescence resonance energy transfer (FRET)-based quantum dot (QD) immunoassay for detection and identification of Aspergillus amstelodami was developed. Biosensors were formed by conjugating QDs to IgG antibodies and incubating with quencher-labeled analytes; QD energy was transferred to the quencher species through FRET, resulting in diminished fluorescence from the QD donor. During a detection event, quencher-labeled analytes are displaced by higher affinity target analytes, creating a detectable fluorescence signal increase from the QD donor. Conjugation and the resulting antibody:QD ratios were characterized with UV-Vis spectroscopy and QuantiT protein assay. The sensitivity of initial fluorescence experiments was compromised by inherent autofluorescence of mold spores, which produced low signal-to-noise and inconsistent readings. Therefore, excitation wavelength, QD, and quencher were adjusted to provide optimal signal-to-noise over spore background. Affinities of anti-Aspergillus antibody for different mold species were estimated with sandwich immunoassays, which identified A. fumigatus and A. amstelodami for use as quencher-labeled- and target-analytes, respectively. The optimized displacement immunoassay detected A. amstelodami concentrations as low as 10(3) spores/mL in five minutes or less. Additionally, baseline fluorescence was produced in the presence of 10(5) CFU/mL heat-killed E. coli O157:H7, demonstrating high specificity. This sensing modality may be useful for identification and detection of other biological threat agents, pending identification of suitable antibodies. Overall, these FRET-based QD-antibody biosensors represent a significant advancement in detection capabilities, offering sensitive and reliable detection of targets with applications in areas from biological terrorism defense to clinical analysis.
| S-EPMC3231422 | biostudies-literature
Project description:The long turnaround time in antimicrobial susceptibility testing (AST) endangers patients and encourages the administration of wide spectrum antibiotics, thus resulting in alarming increases of multidrug resistant pathogens. A method for faster detection of bacterial proliferation presents one avenue toward addressing this global concern. We report on a label-free asynchronous magnetic bead rotation (AMBR) based viscometry method that rapidly detects bacterial growth and determines drug sensitivity by measuring changes in the suspension's viscosity. With this platform, we observed the growth of a uropathogenic Escherichia coli isolate, with an initial concentration of 50 cells per drop, within 20 min; in addition, we determined the gentamicin minimum inhibitory concentration (MIC) of the E. coli isolate within 100 min. We thus demonstrated a label-free, microviscometer platform that can measure bacterial growth and drug susceptibility more rapidly, with lower initial bacterial counts than existing commercial systems, and potentially with any microbial strains.
| S-EPMC3381929 | biostudies-literature
Project description:Designing and implementing means of locally trapping magnetic beads and understanding the factors underlying the bead capture force are important steps toward advancing the capture-release process of magnetic particles for biological applications. In particular, capturing magnetically labeled cells using magnetic microstructures with perpendicular magnetic anisotropy (PMA) will enable an approach to cell manipulation for emerging lab-on-a-chip devices. Here, a Co (0.2 nm)/Ni (0.4 nm) multilayered structure was designed to exhibit strong PMA and large saturation magnetization (Ms ). Finite element simulations were performed to assess the dependence of the capture force on the value of Ms . The simulated force profile indicated the largest force at the perimeter of the disks. Arrays of Co/Ni disk structures of (4-7) μm diameter were fabricated and tested in a microchannel with suspended fluorescent magnetic beads. The magnetic beads were captured and localized to the edge of the disks as predicted by the simulations. This approach has been demonstrated to enable uniform assembly of magnetic beads without external fields and may provide a pathway toward precise cell manipulation methods.
| S-EPMC7552876 | biostudies-literature
Project description:Detection of hepatitis B Virus surface antigen (HBsAg) is an established method for diagnosing both acute and chronic hepatitis B virus (HBV) infection. In addition to enzyme immunoassays (EIAs), rapid diagnostic tests (RDTs) are available for the detection of HBsAg in resource-poor settings. However, the available RDTs have inadequate sensitivity and therefore are not suitable for diagnosis of patients with low levels of HBsAg and for blood screening. To provide a high-sensitivity RDT, we developed a lateral flow immunoassay (LFIA) for HBsAg utilizing upconverting nanoparticle (UCNP) reporter. The UCNP-LFIA can use whole blood, serum, or plasma and the results can be read in 30 min using a reader device. When compared with a commercial conventional visually read LFIA, the developed UCNP-LFIA had a Limit of Detection (LoD) of 0.1 IU HBsAg/ml in spiked serum, whereas the LoD of the conventional LFIA was 3.2 IU HBsAg/ml. The developed UCNP-LFIA fulfills the WHO criterion for blood screening (LoD ≤ 0.13 IU HBsAg/ml) in terms of LoD. The UCNP-LFIA and conventional LFIA were evaluated with well-characterized sample panels. The UCNP-LFIA detected 20/24 HBsAg-positive samples within the HBsAg Performance Panel and 8/10 samples within the Mixed Titer Performance Panel, whereas the conventional LFIA detected 8/24 and 4/10 samples in these panels, respectively. The performance of the assays was further evaluated with HBsAg-positive (n = 108) and HBsAg-negative (n = 315) patient samples. In comparison with a central laboratory test, UCNP-LFIA showed 95.4% (95% CI: 89.5-98.5%) sensitivity whereas sensitivity of the conventional LFIA was 87.7% (95%CI: 79.9-93.3%).
| S-EPMC7813740 | biostudies-literature
Project description:The field of synthetic microswimmers, micro-robots moving in aqueous environments, has evolved significantly in the last years. Micro-robots actuated and steered by external magnetic fields are of particular interest because of the biocompatibility of this energy source and the possibility of remote control, features suited for biomedical applications. While initial work has mostly focused on helical shapes, the design space under consideration has widened considerably with recent works, opening up new possibilities for optimization of propellers to meet specific requirements. Understanding the relation between shape on the one hand and targeted actuation and steerability on the other hand requires an understanding of their propulsion behavior. Here we propose hydrodynamic simulations for the characterization of rigid micropropellers of any shape, actuated by rotating external magnetic fields. The method consists of approximating the propellers by rigid clusters of spheres. We characterize the influence of model parameters on the swimming behavior to identify optimal simulation parameters using helical propellers as a test system. We then explore the behavior of randomly shaped propellers that were recently characterized experimentally. The simulations show that the orientation of the magnetic moment with respect to the propeller's internal coordinate system has a strong impact on the propulsion behavior and has to be known with a precision of ≤ 5° to predict the propeller's velocity-frequency curve. This result emphasizes the importance of the magnetic properties of the micropropellers for the design of desired functionalities for potential biomedical applications, and in particular the importance of their orientation within the propeller's structure.
| S-EPMC7805997 | biostudies-literature
Project description:UnlabelledMagnetic manganese ferrite (MnFe2O4) nanoparticles with approximately 100nm in diameter were used to improve the performance of an immunoassay for detecting influenza infections. The synthesized nanoparticles were tested for long-term storage to confirm the stability of their thermal decomposition process. Then, an integrated microfluidic system was developed to perform the diagnosis process automatically, including virus purification and detection. To apply these nanoparticles for influenza diagnosis, a micromixer was optimized to reduce the dead volume within the microfluidic chip. Furthermore, the mixing index of the micromixer could achieve as high as 97% in 2seconds. The optical signals showed that this nanoparticle-based immunoassay with dynamic mixing could successfully achieve a detection limit of influenza as low as 0.007 HAU. When compared with the 4.5-?m magnetic beads, the optical signals of the MnFe2O4 nanoparticles were twice as sensitive. Furthermore, five clinical specimens were tested to verify the usability of the developed system.From the clinical editorIn this study, magnetic manganese ferrite nanoparticles were used to improve the performance of a novel immunoassay for the rapid and efficient detection of influenza infections.
| S-EPMC7106285 | biostudies-literature