A novel colorimetric competitive aptamer assay for lysozyme detection based on superparamagnetic nanobeads (original) (raw)
Related papers
Label-free detection of lysozyme in wines using an aptamer based biosensor and SPR detection
Sensors and Actuators B: Chemical, 2015
The optimization of an aptasensor with SPR detection allowed the determination of an allergen protein, lysozyme, with high accuracy, good sensitivity and a detection limit of 2.4 nM for spiked red and white wines. The interaction of lysozyme with specific phenolic compounds from wine was evidenced by circular dichroism and SPR. Quantitative recovery factors higher than 88% for wine lysozyme were achieved based on a sample pre-treatment procedure with high concentration of sodium chloride and Tween-20. This aptasensor is useful to monitor the lysozyme levels during winemaking. Also reported is a new chromatographic method based on size exclusion that provided a good separation of lysozyme from wine components, with a detection limit of 35 nM or 0.5 g/mL.
Bioelectrochemistry, 2015
This work presents a comparison of two different aptamers (COX and TRAN) for the detection of the ubiquitous protein lysozyme using aptamer-based biosensors. The detection is based on the specific recognition by the aptamer immobilized on screen printed carbon electrodes (SPCEs) via diazonium coupling reaction. The quantitative detection of lysozyme protein was achieved by electrochemical impedance spectroscopy (EIS). Very good linear ranges and detection limits for the lysozyme detection were obtained, from 0.025 to 1 µM and 725nM using aptamer COX and from 0.025 to 1 µM and 31.7nM using aptamer TRAN. The obtained results showed that the developed aptasensors exhibit good specificity, stability and reproducibility for lysozyme detection. The aptasensors were also tested in wine samples; very good recovery rates were obtained in the range from 96.4 to 102% for lysozyme detection. The recovery rates confirm the reliability and suitability of the developed method in wine matrix. The developed method could be a useful and promising platform for detection of lysozyme in different applications.
Aptamer-Based Electrochemical Sensing of Lysozyme
Chemosensors, 2016
Protein analysis and quantification are required daily by thousands of laboratories worldwide for activities ranging from protein characterization to clinical diagnostics. Multiple factors have to be considered when selecting the best detection and quantification assay, including the amount of protein available, its concentration, the presence of interfering molecules, as well as costs and rapidity. This is also the case for lysozyme, a 14.3-kDa protein ubiquitously present in many organisms, that has been identified with a variety of functions: antibacterial activity, a biomarker of several serious medical conditions, a potential allergen in foods or a model of amyloid-type protein aggregation. Since the design of the first lysozyme aptamer in 2001, lysozyme became one of the most intensively-investigated biological target analytes for the design of novel biosensing concepts, particularly with regards to electrochemical aptasensors. In this review, we discuss the state of the art of aptamer-based electrochemical sensing of lysozyme, with emphasis on sensing in serum and real samples.
Label-free electrochemical aptasensor for the detection of lysozyme
Talanta, 2009
This work reports the advantages of a label free electrochemical aptasensor for the detection of lysozyme. The biorecognition platform was obtained by the adsorption of the aptamer on the surface of a carbon paste electrode (CPE) previously blocked with mouse immunoglobulin under controlled-potential conditions. The recognition event was detected from the decrease in the guanine and adenine electro-oxidation signals produced as a consequence of the molecular interaction between the aptamer and lysozyme. The biosensing platform demonstrated to be highly selective even in the presence of large excess (9-fold) of bovine serum albumin, cytochrome C and myoglobin. The reproducibility for 10 repetitive determinations of 10.0 mg L −1 lysozyme solution was 5.1% and 6.8% for guanine and adenine electro-oxidation signals, respectively. The detection limits of the aptasensor were 36.0 nmol L −1 (if considering guanine signal) and 18.0 nmol L −1 (if taking adenine oxidation current). This new sensing approach represents an interesting and promising alternative for the electrochemical quantification of lysozyme.
Analytical Chemistry, 2011
Magnetic relaxation switch (MRSw) detection is based on aggregate formation or dissociation when magnetic nanoparticles (MNPs) bind to target molecules. In the aggregated state, the dephasing rate of nearby proton spins is higher than in the dispersed state, resulting in a decrease in the spin-spin relaxation time, T 2 . In this work, an MRSw-based nanosensor for lysozyme (Lys) protein detection was achieved using iron oxide nanoparticles conjugated with either Lys aptamer or linker DNA, which can hybridize with the extended part of the aptamer to form clusters. Upon the addition of Lys, the aptamers bind with their targets, leading to disassembly of clusters and an increase in T 2 . A detection limit in the nanomolar range was achieved for Lys detection in both buffer and human serum. The determination of Lys level in different types of cancer cell lysates was also performed to demonstrate detection in real clinical samples.
2018
Foodborne illness is a major concern worldwide due to its impacts towards health, economics and society. One of the major foodborne diseases is salmonellosis that is caused by the members of the species Salmonella enterica. Even though culture method is the gold standard for pathogen detection, this method is too time-consuming and laborious. Therefore, many alternative methods have been developed to increase sensitivity, specificity and speed of detection. Foodborne pathogens detection involves various aspects which are sample preparation, isolation and detection. The objectives of the study were to improve the food sample preparation by using DNA aptamer magnetic separation (AMS) and evaluate the sensitivity and specificity of an aptamer-based biosensor (aptasensor) for Salmonella detection. In the first part of this study, biotinylated aptamer was coupled with the streptavidin magnetic beads and then used to isolate whole Salmonella cells, followed by detection using PCR targetin...
Electroanalysis, 2009
A magnetic particle assay has been designed herein that can report the interactions of DNA aptamers with their cognate protein targets lysozyme (LYS) and human thrombin (THR). Electrochemical sensing of the biomolecular recognition between each aptamer and its target was explored by using a disposable graphite electrode, PGE, in combination with differential pulse voltammetry (DPV). The magnitudes of the oxidation signals of LYS and THR were measured at þ 780 mV and þ 680 mV, respectively, after interaction with the cognate aptamers attached to the surface of magnetic particles. The detection limits estimated for signal to noise ratios above 3.0 correspond to the concentrations of 10.77 mg/mL LYS (769 nM) and 2.00 mg/mL THR (54.5 nM). Our aptamer based approach that combines magnetic particles with a disposable graphite electrode performs well compared to other aptamer-based sensor-formats for quantitative protein detection with respect to sensitivity, selectivity, detection limit, and reproducibility.
BioTechniques, 2002
Systematic evolution of ligands by exponential enrichment (SELEX) was used to develop DNA ligands (aptamers) to cholera whole toxin and staphylococcal enterotoxin B (SEB). Affinity selection of aptamers was accomplished by conjugating the biotoxins to tosyl-activated magnetic beads. The use of magnetic beads reduces the volumes needed to perform aptamer selection, thus obviating alcohol precipitation and allowing direct PCR amplification from the bead surface. Following five rounds of SELEX, 5′-biotinylated aptamers were bound to streptavidin-coated magnetic beads and used for the detection of ruthenium trisbypyridine [Ru(bpy)32+]-labeled cholera toxin and SEB by an electrochemiluminescence methodology. A comparison of control (double-stranded) aptamer binding was made with aptamers that were heat denatured at 96°C (single-stranded) and allowed to cool (conform) in the presence of biotoxin-conjugated magnetic beads. Results suggest that control aptamers performed equally well when c...