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The rapid and cost-effective detection of bacteria is of great importance to ensuring food safety, preventing food poisoning. Herein, we developed a sensitive detection of Escherichia coli (E. coli) using bacteria-specific aptamer in conjunction with microchip capillary electrophoresis-coupled laser-induced fluorescence (MCE-LIF). Based on the differences between charge to mass ratios of free aptamer and bacteria-aptamer complex, which influence their electrophoretic mobilities, the separation of free aptamers and complex peaks by MCE could be achieved. Under optimal conditions, the sensitive detection of E. coli was achieved with a detection limit of 3.7 × 102 CFU mL−1, at a fast response of 135 s and a short detection length of 2.3 cm. The spiked recovery experiment showed that E. coli could be recovered from spiked drinking water and milk samples with recovery rates of 94.7% and 92.8%, respectively. This work demonstrates that the established detection strategy can be a useful tool for the detection and/or monitoring of E. coli in food and environment.

Abnormal glycosylation of exosomal proteins is related to many diseases. However, there is still a lack of convenient and easy methods for the determination of exosomal glycoproteins. In this work, a ratiometric electrochemical sensor based on the recognition of glycoproteins by boronic acid and core-shell nanoparticles of silica-silver (SiO2@Ag) amplified signals was developed for the highly sensitive detection of exosomal glycoproteins. The CD63 aptamer-SiO2-N-(2-((2-aminoethyl)disulfanyl)ethyl) ferrocene carboxamide (FcNHSSNH2) probe was first connected to graphene oxide-cucurbit [7] (GO-CB [7]) modified GCE through host-guest recognition. The CD63 aptamer was employed for the specific capture of exosomes, and the FcNHSSNH2 molecule was used as the internal reference signal of the sensor. The mercaptophenylboronic acid (MPBA) of MPBA-SiO2@Ag probe was used for the identification of exosomes surface glycoproteins. SiO2 nanoparticle has a large specific surface area, which can load a large amount of silver nanoparticles (AgNPs) for electrochemical signal amplification. The results were expressed as the current ratio of AgNPs and FcNHSSNH2. The introduction of the internal reference molecule FcNHSSNH2 could effectively reduce the measurement error caused by the different DNA density of the substrate, and further improve the sensitivity and accuracy of the detection. Under the optimal experimental conditions, this sensor allowed the sensitive detection of exosomal glycoproteins in the range of 4.2 × 102 to 4.2 × 108 particles/μL with a limit of detection (LOD) of 368 particles/μL. Furthermore, the ratiometric electrochemical sensor could be employed for the detection of exosomal glycoproteins in human serum samples, which has a good clinical application prospect.

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the main pathogens involved in hospital and community infection. To rapidly and sensitively detect the mecA gene, which is relevant to methicillin-resistant strains, microchip electrophoresis (MCE) integrated with isothermal strand-displacement polymerase reaction (ISDPR) was developed. In the ISDPR signal recycle amplification, the target DNA opened the DNA hairpin structure by specifically binding with the hairpin probe (HP), and then the primer hybridized with the probe and released the target DNA in the presence of Klenow Fragment exo− (KF exo−) polymerase. The released target DNA hybridized with the next HP and then was displaced by the primer again, consequently achieving target recycling and amplification. The amplified products of the HP-cDNA duplex were separated rapidly from other DNAs by MCE. Under optimal conditions, the limit of detection of the target DNA was as low as 12.3 pM (S/N = 3). The proposed ISDPR with MCE method was also successfully applied to detect methicillin-resistant S. aureus, and the experimental results showed that it had some advantages such as being label free, ultrasensitive, rapid and well separated.

Simultaneous detection of various intracellular biomarkers is promising for early diagnosis and treatment of cancer. Herein, a split primer ligation-triggered catalyzed hairpin assembly-based on dual-signal electrochemical biosensor was constructed for the determination of two pairs of cancer mRNAs: TK1 and c-myc, survivin and GalNAc-T by using ferrocene molecular beacon and hemin molecular beacon as detection signal sources. Each pair of targets exists simultaneously, can release the split primers and ligated as the integral primers, hybridization occurred between the integral primers and part of MBs, causing a double-stranded DNA formed. The probes hybridized with the unfolded MBs and displaced integral primers. Finally, the displaced integral primers again hybridized with the MBs and initiated cycle amplification. Under the optimal conditions, the detection limit of TK1 and c-myc mRNA is as low as 0.022 nM, and that of survivin and GalNAc-T mRNA is 0.029 nM. In addition, two pairs of cancer mRNAs could act as outputs to activate an AND logic gate.

The separation of chiral amino acids using microchip electrophoresis (MCE) was investigated using chiral nematic mesoporous silica (CNMS) as the chiral stationary phase, with hydroxypropyl-β-cyclodextrin (HP-β-CD) as the chiral selector. Individually, neither CNMS nor HP-β-CD achieved separation, so they were combined. Ten chiral amino acids (phenylalanine, tryptophan, glutamic, alanine, serine, aspartic acid, cysteine, methionine, tyrosine, and histidine) were selected as the model analytes. Under optimized conditions, we achieved baseline separation of six chiral amino acids, and the other four chiral amino acids displayed improved resolution. These results indicate the presence of a synergistic effect between CNMS and HP-β-CD, showing that the combination of a chiral stationary phase and a chiral additive is a promising approach for enantioseparation using MCE.

A capillary column was prepared by continuously packing ostadecylsilane (ODS) and taurine derivatized silica (TDS) in one column without interface. This continuously packing chromatography (CPC) column is easy to operate, has good stability and shows simultaneously separation of both polar and non-polar compounds. The simultaneous separation of a series of complex samples with highly hydrophobic components (benzene, toluene, ethylbenzene, and PAHs) and highly hydrophilic components (biogenic amines, bases and nucleosides) using this CPC method was investigated. The relative parameters such as the volume fraction of acetonitrile and length of the ODS and the TDS phases were investigated and optimized. The experimental results show that this column combines the advantages of both ODS and TDS stationary phases, and exhibits a reversed phase liquid chromatography (RPLC) mode followed by a hydrophilic interaction liquid chromatography (HILIC) mode when 80% of acetonitrile was used in the mobile phase. The satisfactory results indicate that the CPC method provides an easy way to simultaneously separate polar and non-polar compounds.

Herein, a novel and convenient electrochemically active-inactive switching molecular beacon based on hemin (Hs-MB) has been designed for easy discrimination of single nucleotide polymorphisms (SNPs) and sensitive detection of insulin. The electrochemically active changing capability of Hs-MB is based on two identical hemin groups labeled at both ends of MB sequence in dimer or monomer forms depending on the conformation of MB which is in stem-loop structure or line shaped structure. The Hs-MB assay permits discrimination of SNPs and the highly sensitive and specific detection of insulin with detection limit successively as low as 0.5 pmol/L. Even at a very low target concentration, the Hs-MB assay also shows a good specificity in the presence of other potentially interfering components. The experimental results also show that Hs-MB can also be used for the accurate and rapid monitoring of insulin secretion by glucose-stimulated from MIN6 cells at different time periods, demonstrating that Hs-MB has potential in monitoring of biomarker variation in vivo.

This paper reports the development of an ultra-sensitive colorimetric method for the detection of trace mercury ions involving DNAzymes, Au nanoparticle aggregation, magnetic nanoparticles and an endonuclease. DNAzyme-sensing elements are conjugated to the surface of Au nanoparticle-2, which can crosslink with the T-rich strands coated on Au nanoparticle-1 to form Au nanoparticle aggregation. Other T-rich stands are immobilized on the surface of MNPs. The specific hybridization of these two T-rich strands depends on the presence of Hg(2+), resulting in the formation of a T-Hg(2+)-T structure. Added endonuclease then digests the hybridized strands, and DNAzyme-modified Au NP aggregation is released, catalysing the conversion of the colourless ABTS into a blue-green product by H2O2-mediated oxidation. The increase in the adsorption spectrum of ABTS(+) at 421 nm is related to the concentration of Hg(2+). This assay was validated by detecting mercury ion concentrations in river water. The colorimetric responses were not significantly altered in the presence of 100-fold excesses of other metal ions such as Zn(2+), Pb(2+), Cd(2+), Mn(2+), Ca(2+) and Ni(2+). The inclusion of both Au NP aggregation and an endonuclease enables the assay to eliminate interference from the magnetic nanoparticles with colorimetric detection, decrease the background and improve the detection sensitivity. The calibration curve of the assay was linear over the range of Hg(2+) concentrations from 1 to 30 nM, and the detection limit was 0.8 nM, which is far lower than the 10 nM US EPA limit for drinking water.

In the paper, a new kind of vitamin B 12 (acquo-cobalamine) chemically modified electrode was fabricated and applied in capillary zone electrophoresis coupled with amperometric detection (CZE-AD) for simultaneous determination of six antioxidants in fruits and vegetables. The catalytic electrochemical properties of the chemically modified electrode could obviously enhance oxidation peak heights responses by about five times to glutathione, ascorbic acid, vanillic acid, chlorogenic acid, salicylic acid, and caffeic acid compared with common carbon disk electrode. Furthermore, the effects of working electrode potential, pH and concentration of running buffer, separation voltage and injection time on CZE-AD were investigated. Under the optimum conditions, the six analytes could be completely separated and detected in a borate–phosphate buffer (pH 8.4) within 15 min. Their linear ranges were from 2.5 × 10 −7 to 1.0 × 10 −4 mol L −1 and the detection limits were as low as 10 −8 mol L −1 magnitude ( S / N = 3). The proposed method has been successfully employed to monitor the six analytes in practical samples with recoveries in the range 96.0–106.0% and RSDs less than 5.0%. Above results demonstrate that capillary zone electrophoresis coupled with electrochemical detection using vitamin B 12 modified electrode as detector is of convenient preparation, high sensitivity, good repeatability, and could be used in the rapid determination of practical samples.

In this protocol, a fluorescent aptasensor based on magnetic separation for simultaneous detection thrombin and lysozyme was proposed. Firstly, one of the anti-thrombin aptamer and the anti-lysozyme aptamer were individually immobilized onto magnetic nanoparticles, acting as the protein captor. The other anti-thrombin aptamer was labeled with rhodamine B and the anti-lysozyme aptamer was labeled with fluorescein, employing as the protein report. By applying the sandwich detection strategy, the fluorescence response at 515 nm and 578 nm were respectively corresponding to lysozyme and thrombin with high selectivity and sensitivities. The fluorescence intensity was individually linear with the concentration of thrombin and lysozyme in the range of 0.13–4 nM and 0.56–12.3 nM, and the detection limits were 0.06 nM of thrombin and 0.2 nM of lysozyme, respectively. The preliminary study on simultaneous detection of thrombin and lysozyme in real plasma samples was also performed. It shows that the proposed approach has the good character for simultaneous multiple protein detection.

Despite the separation efficiency of capillary electrophoresis (CE) is much higher than other chromatographic methods, it is sometimes difficult to perfectly separate the complex ingredients in biological samples. One possible and simple way to develop the separation effect in CE is to add some modifiers in the running buffer. In this paper, the suitable running buffer modifiers were explored to simultaneously separate and detect six typical flavonoids (apigenin, luteolin, kaempferol, quercetin, (+)-catechin and (-)-epicatechin) which are the main active ingredients in chrysanthemum by capillary zone electrophoresis with amperometric detection (CZE-AD). It was found that when beta-cyclodextrin (beta-CD) and the mixture of methanol and ethanol were used as running buffer modifiers, a baseline separation of the six analytes could be accomplished in less than 20 min and the detection limits were as low as 10(-7) or 10(-8)gm l(-1). Other factors affecting the CZE separation, such as working potential, pH value and ionic strength of running buffer, separation voltage and sample injection time were extensively investigated. Under the optimum conditions, a successful practical application on the determination of chrysanthemum samples confirmed the validity and practicability of this method.

In this paper, a polyamide-modified carbon paste electrode in capillary zone electrophoresis with amperometric detection (CZE–AD) was firstly applied to the determination of four carbamate pesticides: fenobucarb, isoprocarb, metolcarb and carbaryl. The four carbamates were hydrolyzed in alkalescent aqueous solutions, resulting in the formation of 2-sec-butylphenol, 2-isopropylphenol, m-cresol and α-naphthol, which could be determined by amperometry after capillary electrophoretic separation. Under the selected optimum conditions, the four analytes could be perfectly separated within 23 min. The linear ranges of 2-sec-butylphenol, 2-isopropylphenol and m-cresol were from 1.0 × 10 −7 to 2.0 × 10 −5 mol L −1 and that of α-naphthol was from 2.0 × 10 −7 to 2.0 × 10 −5 mol L −1 and their detection limits were 3.0 × 10 −8 , 3.0 × 10 −8 , 3.0 × 10 −8 and 6.0 × 10 −8 mol L −1 , respectively ( S / N = 3). Fenobucarb, isoprocarb, metolcarb and carbaryl can be indirectly determined by this CZE–AD method with recovery of 105, 104, 110 and 98% and R.S.D. of 4, 3, 4 and 3%, respectively. Above results demonstrated that this method was of high sensitivity, good repeatability and could be used in the rapid determination of the pesticide residues.

In this paper, capillary zone electrophoresis with amperometric detection (CZE–AD) was firstly applied to the simultaneous separation and determination of nitroaniline positional isomers. The three analytes could be perfectly analyzed by using the buffer of extreme pH. The effects of several important factors were investigated to find optimum conditions. A carbon-disk electrode was used as working electrode. The optimal conditions were 40 mmol/L tartaric acid–sodium tartrate (pH 1.2) as running buffer, 17 kV as separation voltage and 1.10 V (versus saturated calomel reference electrode, SCE) as detection potential. Under the optimum conditions, o-, m- and p-nitroaniline were separated successfully and good linearity, reproducibility and recovery results were obtained. The detection limit for m-nitroaniline was as low as at 9.06 × 10−9 mol/L. This proposed method demonstrated long-term stability and reproducibility with relative standard deviations of less than 1.8% for migration time and 1.1% for peak areas. The utility of this method was demonstrated by monitoring dyestuff wastewater and the assay results were satisfactory.

A synthesized 24-mer single-stranded deoxyribonucleic acid (ssDNA) was covalently immobilized onto a self-assembled aminoethanethiol monolayer modified gold electrode, using water-soluble 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide (EDC). The covalently immobilized ssDNAs were hybridized with complementary ssDNA (cDNA) or yAL 3 gene in solution, forming double-stranded DNAs (dsDNA). Meanwhile, daunomycin as an electrochemical active intercalator in the hybridization buffer solution was intercalated into the dsDNA to form a dsDNA/daunomycin system on the gold electrode surface, which was used for DNA electrochemical sensor. The cathodic waves of daunomycin bound to the double-stranded DNA (dsDNA) by linear sweep voltammetry were utilized to detect the cDNA. The cathodic peak current ( i pc ) of duanomycin was linearly related to the concentrations of cDNA between 0.1 μg ml −1 and 0.1 ng ml −1 . The detection limit was about 30 pg ml −1 .

A sensitive and label-free electrochemiluminescence (ECL) aptasensor for the detection of adenosine triphosphate (ATP) was successfully designed using host–guest recognition between a metallocyclodextrin complex, i.e., tris(bipyridine)ruthenium(II)-β-cyclodextrin [tris(bpyRu)-β-CD], and an ATP-binding aptamer. In the protocol, the NH2-terminated aptamer was immobilized on a glassy carbon electrode (GCE) by a coupling interaction. After host–guest recognition between tris(bpyRu)-β-CD and aptamer, the tris(bpyRu)-β-CD/aptamer/GCE produced a strong ECL signal as a result of the photoactive properties of tris(bpyRu)-β-CD. However, in the presence of ATP, the ATP/aptamer complex was formed preferentially, which restricted host–guest recognition, and therefore less tris(bpyRu)-β-CD was attached to the GCE surface, resulting in an obvious decrease in the ECL intensity. Under optimal determination conditions, an excellent logarithmic linear relationship between the ECL decrease and ATP concentration was obtained in the range 10.0–0.05 nM, with a detection limit of 0.01 nM at the S/N ratio of 3. The proposed ECL-based ATP aptasensor exhibited high sensitivity and selectivity, without time-consuming signal-labeling procedures, and is considered to be a promising model for detection of aptamer-specific targets.

As a kinetic process, chemiluminescence (CL) met great challenges while it was used in the detection methods coupled with capillary electrophoresis (CE). In this investigation, a newly recorded ultra-fast CL reaction of luminol–KIO4–K3Fe(CN)6 was observed to be completed in 0.65 s. It was adopted in a simple CE-CL system efficiently to avoid the peak-tailing and overlapping. With this compact system, an indirect determination of rhamnose, d -fructose, sucrose and β-cyclodextrin was realized based on the corresponding negative CL peaks. These peaks were due to the displacement of luminol anions by the analyzed saccharide molecules in alkaline separation electrolyte. In this way, these four saccharides could be separated and determined in 16 min with adequate sensitivities and stabilities. No derivatization or pretreatment was required for the analysis, and it presents an attractive opportunity for routine tests of mono-, di- and oligo-saccharides in a compact CE-CL system, even as a microchip device.

A solid-state electrochemiluminescence (ECL) biosensing switch based on special ferrocene-labeled molecular beacon (Fc-MB) has been successfully developed for T4 DNA ligase detection. Such special switch system consisted of two main parts, an ECL substrate and an ECL intensity switch. The ECL substrate was made by modifying the complex of Au nanoparticle and Ruthenium (II) tris-(bipyridine) (Ru(bpy)(3)(2+)-AuNPs) onto Au electrode. A molecular beacon labeled by ferrocene as the ECL intensity switch. The molecular beacon is designed with special base sequence, which could combine with its target biomolecule via the reaction of the repair and recombination of nucleic acids by DNA ligase. During the reaction, the molecular beacon opened its stem-loop, and the labeled Fc was consequently kept away from the ECL substrate. Such structural change resulted in an obvious increment in ECL intensity due to the decreased Fc quenching effect to the ECL substrate. The analysis results are sensitive and specific.

An aptamer-based assay for thrombin with high specificity and sensitivity was presented. In the protocol, the aptamer for thrombin was immobilized on magnetic nanoparticle, and its complementary oligonucleotide was labeled with gold nanoparticles, then the aptamer was hybridized with the complementary oligonucleotide to form the duplex structure as a probe, this probe could be used for the specific recognition for thrombin. In the presence of thrombin, the aptamer prefer to form the G-quarter structure with thrombin, resulting in the dissociation of the duplex of the probe and the release of the gold labeled oligonucleotide. Upon this, we were able to detect thrombin through the detection of the electrochemical signal of gold nanoparticles. The strategy combines with the high specificity of aptamer and the excellent characteristics of nanoparticles. This assay is simple, rapid, sensitive and highly specific, it does not require labeling of thrombin, and it could be applied to detect thrombin in complex real sample. The method shows great potential in other protein analysis and in disease diagnosis.

A simple, sensitive and selective flow injection-chemiluminescence method for the determination of sulfadiazine in compound naristillae has been investigated. It is based upon the chemilimunescence reaction of sulfadiazine, formaldehyde and potassium permanganate in polyphosphate acid medium. The optimum conditions for the chemiluminescence emission were investigated. Under the optimum conditions, the linear range for the determination of sulfadiazine was 8.0x10(-7) to 2.0x10(-4)mol/L with a detection limit of 2.0x10(-7)mol/L calculated as proposed by IUPAC and a relative standard deviation of 2.53% for 11 solutions of 5.0x10(-5)mol/L sulfadiazine on the same day. It was also found that the coexisting ephedrine hydrochloride did not interfere with this determination. This led to the successful application of the proposed method for the direct and selective determination of sulfadiazine in compound naristillae.

Based on self-assembled biotinylated disulfide derivative monolayer on gold electrode, the sensors immobilized monolayer or multilayer membranes composed of avidin and biotinlabeled glucose oxidase (B·GOD) or of avidin-B·GOD complex (ABC) and B·COD were prepared. The present technique may be useful for controlling the enzyme content of the sensors in molecular level by repeating the deposition of enzyme layers. The sensors have the characteristics of shorter response time, higher sensitivity. The linear range is from 6.0 × 10−6 − 5.0 × 10−3 M. The sensor can be used for more than 1 month and can be reactivated. The sensor was used to determine glucose in human blood serum, and the results are satisfactory.