Photoinduced Electron Transfer Mediated by π-Stacked Thymine−Hg²⁺−Thymine Base Pairs

Fluorescence spectra of thymine-containing oligonucleotides labeled with donor/acceptor in the presence of Hg2+ ions were investigated. Formation of T−Hg2+−T base pairs gives rise to a hairpin structure and makes both termini of the oligonucleotide close to each other. For fluorescein or tetramethylrhodamine single-labeled oligonucleotide, fluorescence quenching was observed on addition of Hg2+ ions. For fluorescein and tetramethylrhodamine double-labeled oligonucleotide, the apparent FRET efficiencies decrease unexpectedly in the presence of Hg2+ ions. The unusual fluorescence quenching in the presence of Hg2+ ions was ascribed to formation of T−Hg2+−T base pairs, which can accept and mediate the electron transfer and provide an additional de-excitation process for the excited state of fluorophores via photoinduced electron transfer

Matrix Interference-Free Method for the Analysis of Small Molecules by Using Negative Ion Laser Desorption/Ionization on Graphene Flakes

This work presents a new approach for the analysis of small molecules with direct negative ion laser desorption/ionization (LDI) on graphene flakes. A series of matrix interference-free mass spectra were obtained for the analysis of a wide range of small molecules including peptides, amino acids, fatty acids, as well as nucleosides and nucleotides. The mixture of analytes and graphene flakes suspension were directly pipetted onto a sample plate for LDI-time-of-flight mass spectrometry (TOFMS) analysis. Deprotonated monomeric species [M−H]− ions were homogeneously obtained on uniform graphene flakes film when negative ion mode was applied. In positive ion mode, the analytes were detected in form of multiple adduct ions such as sodium adduct [M+Na]+, potassium adduct [M+K]+, double sodium adduct [M+2Na−H]+, double potassium adduct [M+2K−H]+, as well as sodium and potassium mixed adduct [M+Na+K−H]+. Better sensitivity and reproducibility were achieved in negative ion mode compared to positive ion mode. It is believed that the new method of matrix interference-free negative ion LDI on graphene flakes may be expanded for LDI-MS analysis of various small molecules

Ultrasensitive Aptamer-Based Multiplexed Electrochemical Detection by Coupling Distinguishable Signal Tags with Catalytic Recycling of DNase I

This work reports an aptamer-based, disposable, and multiplexed sensing platform for simultaneous electrochemical determination of small molecules, employing adenosine triphosphate (ATP) and cocaine as the model target analytes. The multiplexed sensing strategy is based on target-induced release of distinguishable redox tag-conjugated aptamers from a magnetic graphene platform. The electronic signal of the aptasensors could be further amplified by coupling DNase I with catalytic recycling of self-produced reactants. The assay was based on the change in the current at the various peak potentials in the presence of the corresponding signal tags. Experimental results revealed that the multiplexed electrochemical aptasensor enabled the simultaneous monitoring of ATP and cocaine in a single run with wide working ranges and low detection limits (LODs: 0.1 pM for ATP and 1.5 pM for cocaine). This concept offers promise for rapid, simple, and cost-effective analysis of biological samples

Carbon Dioxide Gas Sensor Based on Ionic Liquid-Induced Electrochemiluminescence

Electrochemiluminescence of the luminol–O2 system in an electrolyte-free N,N-dimethylformamide (DMF)–dipropylamine (DPA) cosolution is induced by the formation of a carbamate ionic liquid (IL) from the reaction between CO2 and DPA, on the basis of which a facile ECL sensor for measuring atmospheric CO2 has been developed. This ECL sensing method shows several advantages in the detection of CO2, such as high safety, high selectivity, wide linear response range, and good sensitivity. The gas sensor was found to have a linear response range from 100 ppm to 100 v/v% and a detection limit of 80 ppm (at signal-to-noise ratio of 3). This is the first reported IL-induced ECL sensor for a gas, thus the principle of this type of sensor and the IL-induced ECL mechanism have been demonstrated in detail

Preparation of Protein-like Silver–Cysteine Hybrid Nanowires and Application in Ultrasensitive Immunoassay of Cancer Biomarker

Novel protein-like silver–cysteine hybrid nanowires (<i>p</i>-SCNWs) have been synthesized by a green, simple, nontemplate, seedless, and one-step aqueous-phase approach. AgNO₃ and l-cysteine were dissolved in distilled water, forming Ag–cysteine precipitates and HNO₃. Under vigorous stirring, the pH of the solution was rapidly adjusted to 9.0 by addition of concentrated sodium hydroxide solution, leading to quick dissolution of the Ag-cysteine precipitates and sudden appearance of white precipitates of <i>p</i>-SCNWs. The <i>p</i>-SCNWs are monodispersed nanowires with diameter of 100 nm and length of tens of micrometers, and have abundant carboxyl (−COOH) and amine (−NH₂) groups at their surfaces, large amounts of peptide-linkages and S-bonding silver ions (Ag⁺) inside, making them look and act like Ag-hybrid protein nanostructures. The abundant −COOH and −NH₂ groups at the surfaces of <i>p</i>-SCNWs have been found to facilitate the reactions between the <i>p</i>-SCNWs and proteins including antibodies. Furthermore, the fact that the <i>p</i>-SCNWs contain large amounts of silver ions enables biofunctionalized <i>p</i>-SCNWs to be excellent signal amplifying chemiluminescence labels for ultrasensitive and highly selective detection of important antigens, such as cancer biomarkers. In this work, the immunoassay of carcinoembryonic antigen (CEA) in human serum was taken as an example to demonstrate the immunoassay applications of antibody-functionalized <i>p</i>-SCNWs. By the novel <i>p</i>-SCNW labels, CEA can be detected in the linear range from 5 to 400 fg/mL with a limit of detection (LOD) of 2.2 fg/mL (at signal-to-noise ratio of 3), which is much lower than that obtained by commercially available enzyme-linked immunosorbent assay (ELISA). Therefore, the synthesized <i>p</i>-SCNWs are envisioned to be an excellent carrier for proteins and related immunoassay strategy would have promising applications in ultrasensitive clinical screening of cancer biomarkers for early diagnostics of cancers

Electrochemical Biosensor for Detection of BCR/ABL Fusion Gene Using Locked Nucleic Acids on 4-Aminobenzenesulfonic Acid-Modified Glassy Carbon Electrode

In this study, an electrochemical DNA biosensor was developed for detection of the breakpoint cluster region gene and the cellular abl (BCR/ABL) fusion gene in chronic myelogenous leukemia by using 18-mer locked, nucleic acid-modified, single-stranded DNA as the capture probe. The capture probe was covalently attached on the sulfonic-terminated aminobenzenesulfonic acid monolayer-modified glassy carbon electrode through the free amines of DNA bases based on the acyl chloride cross-linking reaction. The covalently immobilized capture probe could selectively hybridize with its target DNA to form double-stranded DNA (dsDNA) on the LNA/4-ABSA/GCE surface. Differential pulse voltammetry was used to monitor the hybridization reaction on the capture probe electrode. The decrease of the peak current of methylene blue, an electroactive indicator, was observed upon hybridization of the probe with the target DNA. The results indicated that, in pH 7.0 Tris-HCl buffer solution, the peak current was linear with the concentration of complementary strand in the range of 1.0 × 10−121.1 × 10−11 M with a detection limit of 9.4 × 10−13 M. This new method demonstrates its excellent specificity for single-base mismatch and complementary dsDNA after hybridization, and this probe has been used for assay of PCR real sample with satisfactory results

Immobilization-Free Programmable Hairpin Probe for Ultrasensitive Electronic Monitoring of Nucleic Acid Based on a Biphasic Reaction Mode

This work designs a novel programmable hairpin probe (PHP) for the immobilization-free electrochemical detection of nucleic acid by coupling polymerase/nicking-induced isothermal signal amplification strategy with a biphasic reaction mode for the first time. The designed PHP (including a target-recognition region, a template sequence for enzymatic reaction and an inactivated <i>anti</i>-streptavidin aptamer) could program multiple isothermal reactions in the solution phase accompanying in situ amplified detectable signal at the electrode surface by the labeled ferrocene tag on the PHP. Upon addition of target analyte into the detection solution, target DNA initially hybridized with the recognition region on the PHP. Replication-induced strand-displacement generated an activated <i>anti</i>-streptavidin aptamer with the assistance of polymerase. Then, the polymerase/nicking enzymes could cleave and polymerize repeatedly the replication product, thus resulting in the formation of numerous template-complementary DNA initiator strands. The released initiator strands could retrigger the programmable hairpin probe to produce a large number of activated <i>anti</i>-streptavidin aptamers, which could be captured by the immobilized streptavidin on the electrode, thus activating the electrical contact between the labeled ferrocene and the electrode. Going after the aptamers, the carried ferrocene could produce the in situ amplified electronic signal within the applied potentials. Under optimal conditions, the electrochemical signal increased with the increasing target DNA concentration in the dynamic range from 5 fM to 10 pM with a detection limit (LOD) of 2.56 fM at the 3<i>s</i>_blank criterion. Importantly, the methodology with high specificity was also validated and evaluated by assaying 6 target DNA-spiked human serum and calf thymus DNA samples, and the recoveries were 95–110%. Further work for the programmable hairpin probe could be even utilized in a real world sample to detect miRNA-21 at femtomol level

Microwave-Assisted ¹⁸O-Labeling of Proteins Catalyzed by Formic Acid

Oxygen exchange may occur at carboxyl groups catalyzed by acid. The reaction, however, takes at least several days at room temperature. The long-time exchanging reaction often prevents its application from protein analysis. In this study, an 18O-labeling method utilizing microwave-assisted acid hydrolysis was developed. After being dissolved in 16O/18O (1:1) water containing 2.5% formic acid, protein samples were exposed to microwave irradiation. LC-MS/MS analysis of the resulted peptide mixtures indicated that oxygen in the carboxyl groups from glutamic acid, aspartic acid, and the C-terminal residues could be efficiently exchanged with 18O within less than 15 min. The rate of back exchange was so slow that no detectable back exchange could be found during the HPLC run

Magneto-Controlled Graphene Immunosensing Platform for Simultaneous Multiplexed Electrochemical Immunoassay Using Distinguishable Signal Tags

A novel flow-through multiplexed immunoassay protocol for simultaneous electrochemical determination of carcinoembryonic (CEA) and alpha-fetoprotein (AFP) in biological fluids was designed using biofunctionalized magnetic graphene nanosheets (MGO) as immunosensing probes and multifunctional nanogold hollow microspheres (GHS) as distinguishable signal tags. The probes were fabricated by means of co-immobilization of primary anti-CEA (Ab1) and anti-AFP (Ab2) antibodies on the Fe3O4 nanoparticle-coated graphene nanosheets (MGO-Ab1,2). The reverse-micelle method was used for the synthesis of distinguishable signal tags by encapsulation of horseradish peroxide (HRP)-thionine and HRP-ferrocene into nanogold hollow microspheres, respectively, which were utilized as labels of the corresponding GHS-Ab1 and GHS-Ab2. A sandwich-type immunoassay format was employed for the online detection of CEA and AFP by coupling a flow-through detection cell with an external magnet. The assay was based on the catalytic reduction of H2O2 at the various peak potentials in the presence of the corresponding mediators. Experimental results revealed that the multiplexed electrochemical immunoassay enabled the simultaneous monitoring of AFP and CEA in a single run with wide working ranges of 0.01–200 ng mL–1 for AFP and 0.01–80 ng mL–1 for CEA. The detection limits (LODs) for both analytes at 1.0 pg mL–1 (at 3sB) were very low. No obvious nonspecific adsorption and cross-talk were observed during a series of analyses to detect target analytes. Intraassay and interassay coefficients of variation were <10%. Importantly, the methodology was evaluated for the analysis of clinical serum specimens, receiving a good correlation between the flow-through multiplexed electrochemical immunoassay and an electrochemiluminescence method as a reference

Encapsulation of Strongly Fluorescent Carbon Quantum Dots in Metal–Organic Frameworks for Enhancing Chemical Sensing

Novel highly fluorescent (FL) metal–organic frameworks (MOFs) have been synthesized by encapsulating branched poly-(ethylenimine)-capped carbon quantum dots (BPEI-CQDs) with a high FL quantum yield into the zeolitic imidazolate framework materials (ZIF-8). The as-synthesized FL-functionalized MOFs not only maintain an excellent FL activity and sensing selectivity derived from BPEI-CQDs but also can strongly and selectively accumulate target analytes due to the adsorption property of MOFs. The selective accumulation effect of MOFs can greatly amplify the sensing signal and specificity of the nanosized FL probe. The obtained BPEI-CQDs/ZIF-8 composites have been used to develop an ultrasensitive and highly selective sensor for Cu²⁺ ion, with a wide response range (2–1000 nM) and a very low detection limit (80 pM), and have been successfully applied in the detection of Cu²⁺ ions in environmental water samples. It is envisioned that various MOFs incorporated with FL nanostructures with high FL quantum yields and excellent selectivity would be designed and synthesized in similar ways and could be applied in sensing target analytes