A sensitive "off-on" electrochemiluminescence DNA sensor based on signal cascade amplification circuit and distance-dependent energy transfer.

A sensitive "off-on" electrochemiluminescence (ECL) DNA sensor was constructed based on Exo III-assisted cascade amplification system. In the cascade amplification circuit, target DNA and Exo III cutting substrate were designed into an inverted T-shaped binding mode to form a stable DNA junction, thus effectively triggering Exo III digestion cycle. During the biosensor assembly process, ferrocene (Fc) and distance-dependent ECL resonance energy transfer (ECL-RET) and surface plasmon resonance (SPR) effects were introduced to regulate the ECL of semiconductor quantum dots (QDs). Carboxylated ZnCdSe/ZnS QDs were used as ECL signal probes and K2S2O8 was coreactant, and the initial cathodic ECL signal of QDs was efficiently quenched through electron and energy transfer with Fc and ECL-RET with Au NPs, leaving the system in "off" state. After the products of cascade amplification were introduced into the electrode surface, the single-stranded DNA modified with Fc was displaced, and the distance between Au NPs and QDs became farther, resulting in a transition from ECL-RET to SPR, and then a significant ECL signal boost was achieved, turning the system into "on" state. The combination of efficient cascade amplification system and sensitive "off-on" ECL signal change mode enabled the biosensing platform to detect target DNA with high selectivity (able to distinguish single-base mutated DNA) and ultra-high sensitivity (limit of detection was 31.67 aM, S/N = 3), providing a new perspective for designing highly sensitive and programmable ECL biosensors.

Combination of Ternary Electrochemiluminescence System of BNQDs/AgMOG-K2S2O8 and Electrochemiluminescence Resonance Energy Transfer Strategy for Ultrasensitive Immunoassay of Amyloid-ß Protein.

In this work, based on boron nitride quantum dots (BNQDs) as energy donors and MnO2@MWCNTs-COOH as energy receptors, we designed an efficient electrochemiluminescence resonance energy transfer (ECL-RET) immunosensor for the detection of amyloid-ß (Aß42) protein, a biomarker of Alzheimer's disease (AD). First, the signal amplification of a ternary ECL system composed of BNQDs (as the ECL emitter), K2S2O8 (as the coreactant), and silver metal-organic gels (AgMOG, as the coreaction accelerator) was realized, and PDDA as stabilizer was added, a strong and stable initial ECL signal was obtained. AgMOG could not only support a large amount of BNQDs and Aß42 capture antibody (Ab1) through Ag-N bond but also exhibit excellent ECL catalytic performance and enhance the luminescent intensity of BNQDs@PDDA-K2S2O8 system. In addition, due to the broad absorption spectrum of MnO2@MWCNTs-COOH and the extensive overlap with the ECL emission spectrum of BNQDs, the quenching probe Ab2-MnO2@MWCNTs-COOH could be introduced into the ternary system through a sandwich immune response. On this basis, the signal on-off ECL immunosensor was constructed to achieve the ultrasensitive detection of Aß42 through signal transformation. Under the optimal conditions, the prepared ECL biosensor manifested a wide linear range (10 fg/mL-100 ng/mL) with a detection limit of 2.89 fg/mL and showed excellent stability, selectivity, and repeatability, which provided an effective strategy for the ultrasensitive detection of biomarkers in clinical analysis.

Room-Temperature Synthesized Iron/Cobalt Metal-Organic Framework Nanosheets with Highly Efficient Catalytic Activity toward Luminol Chemiluminescence Reaction.

Two-dimensional (2D) iron/cobalt metal-organic framework nanosheets (Fe/Co-MOF NSs) were synthesized via the cooperative self-assembly reaction of Fe3+/Co2+ and terephthalic acid at room temperature. The as-prepared 2D Fe/Co-MOF NSs display superior performance in catalysis of the chemiluminescence (CL) reaction between luminol and H2O2. The CL spectrum, UV-vis absorption spectroscopy, radical scavenger experiments, and electron spin resonance (ESR) spectroscopy are utilized to research the possible CL mechanism of the luminol-H2O2-Fe/Co-MOF NSs system. All results indicate that Fe/Co-MOF NSs present outstanding peroxidase-like activity and could catalyze H2O2 to produce 1O2, O2·-, and ·OH, which could react rapidly with the luminol anion radical and result in strong CL. With the highly efficient CL of the luminol-H2O2-Fe/Co-MOF NSs system, a sensitive sensor for the detection of dopamine (DA) is developed based on the inhibitory effect of DA on the CL intensity. Good linearity over the range of 50-800 nM is achieved with a limit of detection of 20.88 nM (S/N = 3). This research demonstrates that 2D Fe/Co-MOF NSs is a highly effective catalyst for luminol CL reaction and has great application potential in the CL field.

Sensitive and Amplification-Free Electrochemiluminescence Biosensor for HPV-16 Detection Based on CRISPR/Cas12a and DNA Tetrahedron Nanostructures.

Rapid and accurate detection of biomarkers was very important for early screening and treatment of diseases. Herein, a sensitive and amplification-free electrochemiluminescence (ECL) biosensor based on CRISPR/Cas12a and DNA tetrahedron nanostructures (TDNs) was constructed. Briefly, 3D TDN was self-assembled on the Au nanoparticle-deposited glassy carbon electrode surface to construct the biosensing interface. The presence of the target would activate the trans-cleavage activity of Cas12a-crRNA duplex to cleave the single-stranded DNA signal probe on the vertex of TDN, causing the Ru(bpy)32+ to fall from the electrode surface and weakened the ECL signal. Thus, the CRISPR/Cas12a system transduced the change of target concentration into an ECL signal enabling the detection of HPV-16. The specific recognition of CRISPR/Cas12a to HPV-16 made the biosensor have good selectivity, while the TDN-modified sensing interface could reduce the cleaving steric resistance and improve the cleaving performance of CRISPR/Cas12a. In addition, the pretreated biosensor could complete sample detection within 100 min with a detection limit of 8.86 fM, indicating that the developed biosensor possesses the potential application prospect for fast and sensitive nucleic acid detection.

Electrochemiluminescence Cascade Amplification Platform for Detection of Dual-microRNA and Operation of Concatenated Logic Circuit.

The detection of multiple biomarkers is of great significance to the accurate diagnosis of diseases. Herein, in this work, we constructed an electrochemiluminescence (ECL) cascade amplification platform for dual acute myocardial infarction (AMI)-related microRNA detection. The Zn2+-dependent DNAzyme digestion reaction initiated by miR-133a and the duplex-specific nuclease (DSN) cleavage circuit initiated by miR-499 were carried out independently to form a fuel hairpin DNA and active initiator strand, respectively, to trigger a hybridization chain reaction, which constituted a two-input-regulated "AND" logic circuit based on single ECL signal output. The use of single signal probe (Ru(bpy)32+) avoided the time-consuming and costly process of multiple signal molecule labeling or modification. The independent operation of the DNAzyme digestion reaction and DSN-assisted target recycling improved the detection efficiency of the system. In addition, the detection of each miRNA had undergone a cascade amplification process, which improved the detection sensitivity for each target. Furthermore, benefitting from the strong complexation of EDTA with Zn2+ and the flexible design of DNA sequences, the two-input "AND" logic gate was extended to a four-input "INHIBIT-AND-INHIBIT" concatenated logic circuit, which broadens the application of the ECL method in logic gates. We anticipate that this cascading amplification strategy can be widely applied in accurate diagnosis of AMI and the construction of ECL-based logic devices.

Virus-like Particles as Nanocarriers for Intracellular Delivery of Biomolecules and Compounds.

Virus-like particles (VLPs) are nanostructures assemble from viral proteins. Besides widely used for vaccine development, VLPs have also been explored as nanocarriers for cargo delivery as they combine the key advantages of viral and non-viral vectors. While it protects cargo molecules from degradation, the VLP has good cell penetrating property to mediate cargo passing the cell membrane and released into cells, making the VLP an ideal tool for intracellular delivery of biomolecules and drugs. Great progresses have been achieved and multiple challenges are still on the way for broad applications of VLP as delivery vectors. Here we summarize current advances and applications in VLP as a delivery vector. Progresses on delivery of different types of biomolecules as well as drugs by VLPs are introduced, and the strategies for cargo packaging are highlighted which is one of the key steps for VLP mediated intracellular delivery. Production and applications of VLPs are also briefly reviewed, with a discussion on future challenges in this rapidly developing field.

Versatile Electrochemiluminescence Biosensing Platform Based on DNA Nanostructures and Catalytic Hairpin Assembly Signal Amplification.

Achieving rapid and highly sensitive detection of biomarkers is crucial for disease diagnosis and treatment. Here, a highly sensitive and versatile dual-amplification electrochemiluminescence (ECL) biosensing platform was constructed for target detection based on DNA nanostructures and catalyzed hairpin assembly (CHA). Specifically, when the target DNA was present, it would hybridize with the auxiliary strands (D1 and D2) to form an I-shaped nanostructure, which in turn triggered the subsequent catalytic hairpin assembly reaction to generate plenty of double-stranded DNA complexes (H1-H2). The resulting double-stranded complex could be trapped on the electrode surface and adsorbed the ECL signal probe Ru(phen)32+.We found that the I-shaped nanostructure-triggered CHA reaction had higher amplification efficiency compared with traditional CHA amplification. Thus, a sensitive "signal-on" ECL biosensor was constructed for target DNA detection with a detection limit of 1.09 fM. Additionally, by combining the binding properties of C-Ag+-C with an elaborately designed "Ag+-helper" probe, the proposed strategy could be immediately utilized for the highly sensitive and selective detection of silver ions, demonstrating the versatility of the developed biosensing platform. This strategy provided a new approach with potential applications in disease diagnosis and environmental monitoring.

Contrary Logic Pair Library, Parity Generator/Checker and Various Concatenated Logic Circuits Engineered by a Label-Free and Immobilization-Free Electrochemiluminescence Resonance Energy Transfer System.

Herein, a label-free and immobilization-free electrochemiluminescence resonance energy transfer (ECL-RET) system based on graphitic carbon nitride nanosheets (GCNNs)/Ru(phen)32+ donor/acceptor pair is developed, in which the ECL-RET is regulated by regulating the diffusivity of Ru(phen)32+ molecules toward the negatively charged GCNNs through logically programmed DNA hybridization reactions. The two optical signals of GCNNs (445 nm) and Ru(phen)32+ (593 nm) show completely opposite changes through the same one-time DNA hybridization reaction. Based on this ECL-RET system, a contrary logic pair (CLP) library, a parity generator/checker system for differentiating the erroneous bits during data transmission, the parity checker to identify the even/odd natural numbers from 0 to 9, and a series of concatenated logic circuits including a six-input logic gate capable of implementing of 64 input combinations for meeting the needs of computational complexity are developed. The ECL-RET-based molecular logic system avoids the time-consuming, costly and inefficient labeling procedures and the laborious processes of immobilization, presenting great potential for building more complicated and advanced logic gates, and providing a refreshing inspiration for the construction of combinatorial logic circuits based on ECL method.

Electrochemiluminescence Biosensor Based on Entropy-Driven Amplification and a Tetrahedral DNA Nanostructure for miRNA-133a Detection.

The early and rapid diagnosis of acute myocardial infarction (AMI) is of great significance to its treatment. Here, we developed an electrochemiluminescence biosensor based on an entropy-driven strand displacement reaction (ETSD) and a tetrahedral DNA nanostructure (TDN) for the detection of the potential AMI biomarker microRNA-133a. In the presence of the target, numerous Ru(bpy)32+-labeled signal probes (SP) were released from the preformed three-strand complexes through the process of ETSD. The ETSD reaction cycle greatly amplified the input signal of the target. The released SP could be captured by the TDN-engineered biosensing interface to generate a strong ECL signal. The rigid structure of TDN could significantly improve the hybridization efficiency. With the assistant of double amplification of TDN and ETSD, the developed biosensor has a good linear response ranging from 1 fM to 1 nM for microRNA-133a, and the detection limit is 0.33 fM. Additionally, the constructed biosensor has excellent repeatability and selectivity, demonstrating that the biosensor possesses a great application prospect in clinical diagnosis.

Electrochemical-Based DNA Logic Devices Regulated by the Diffusion and Intercalation of Electroactive Dyes.

Electrochemical-based logic gates are simple to operate, sensitive, controllable, and easy to integrate with silicon-based semiconductor logic devices, showing great application prospects and remaining largely unexplored. Herein, an immobilization-free dual-output electrochemical molecular logic system based on the different diffusivity of electroactive dyes ferrocene (Fc) and methylene blue (MB) toward an indium tin oxide (ITO) electrode under different DNA hybridization reactions was developed. In this system, the hybridization of the catalytic strand IN1 with Fc-modified hairpin DNA H1 triggered an exonuclease III (Exo III) cleavage cycle to obtain free Fc and produce a large number of long double-stranded DNAs via the hybridization chain reaction for intercalating MB, which was previously in the free state. Such a hybridization reaction caused a significant change in the diffusion capacity of MB and Fc toward the ITO electrode, resulting in two electrochemical signals with opposite changes. On this basis, a contrary logic pair library, a parity generator/checker system for differentiating the erroneous bits during data transmission, a parity checker to identify the even/odd natural numbers from 0 to 9, and a series of concatenated logic circuits for meeting the needs of computational complexity were developed. The proposed electrochemical-based molecular logic system greatly expanded the application of the electrochemical method in the construction of logic circuits and provided a conceptual prototype for the development of more advanced and complicated logic devices.

Label-free immunosensor for cardiac troponin I detection based on aggregation-induced electrochemiluminescence of a distyrylarylene derivative.

Herein, the aggregation-induced electrochemiluminescence (AIECL) of a distyrylarylene derivative, 4,4'-bis(2,2-diphenylvinyl)-1,1'-biphenyl (DPVBi), was investigated for the first time. This luminophore exhibits significantly enhanced photoluminescence (PL) and electrochemiluminescence (ECL) emission with the increases of water content in organic/water mixtures. This high luminescence efficiency of DPVBi in aggregate state is due to the fact that the aggregates can reduce the energy loss by restricting the intramolecular motions. The ECL behavior of DPVBi in acetonitrile was investigated by ECL transients and so-called "half-scan" technology, where singlet-singlet annihilation ECL was generated under continuous potential switching. The DPVBi nanobulks (DPVBi NBs) were prepared to improve its application in aqueous media, which could be conveniently cast on electrode surface for developing sensing platform due to its good film-forming nature. The constructed heterogeneous AIECL platform can produce reductive-oxidative and oxidative-reductive ECL by using trimethylamine (TEA) and potassium peroxodisulfate (K2S2O8) as coreactant. On the basis of the higher ECL efficiency of DPVBi NBs/TEA system, a label free immunosensor for cardiac troponin I (cTnI) was developed with the assistance of electrodeposited gold nanoparticles, and it showed a wide linear range of 20 ng/mL~100 fg/mL and low detection limit of 43 fg/mL. Moreover, the constructed immunosensor also exhibited good specificity, stability and satisfied performance in practical sample analysis.

A ratiometric electrochemiluminescence strategy based on two-dimensional nanomaterial-nucleic acid interactions for biosensing and logic gates operation.

Two-dimensional (2D) nanomaterial-nucleic acid interactions have been widely used in the construction of fluorescent sensors, but they are rarely used in the construction of electrochemiluminescent (ECL) sensors and have never been used in the design of ratiometric ECL sensors. Therefore, a ratiometric ECL sensing platform was developed in this study based on the ECL resonance energy transfer (ECL-RET) of graphitic carbon nitride nanosheets (GCNNs)/Ru(bpy)32+ donor/acceptor pair. The 2D GCNNs showed much weaker affinity to the long dsDNA duplexes formed by hybridization chain reaction (HCR) than Ru(bpy)32+-lableled fuel hairpin DNAs (H1 and H2) for HCR. Therefore, the target-initiated HCR resulted in the luminescence enhancement of the GCNNs at 460 nm and the luminescence attenuation of the Ru(bpy)32+ at 610 nm. By measuring the I460 nm/I610 nm ratios, quantitative analysis of microRNA-133a was realized with a limit of detection of 0.41 pM. In addition, this ECL-RET sensing platform can be easily extended to detect metal ions or aptamer substrates by simply redesigning helper DNAs without changing the sequences of fuel hairpin DNAs. Moreover, due to the programmability of HCR, a series of sensitive logic gates ("OR", "INHIBIT", "AND", "NAND" and "INHIBIT-OR") based on the ECL-RET ratiometry can be constructed and responded to as low as 100 pM of Hg2+ or Ag+.

Sensitive and Programmable "Signal-Off" Electrochemiluminescence Sensing Platform Based on Cascade Amplification and Multiple Quenching Mechanisms.

A versatile and sensitive quantum dot (QD)-based "signal-off" electrochemiluminescence (ECL) sensing system was constructed using target-initiated dual Mg2+-dependent DNAzyme (MNAzyme) recycling and catalytic hairpin assembly (CHA) amplification strategies. After the cascade amplification, numerous ferrocene-labeled Y-shaped DNA complexes generated on the QD-modified electrode surface. In the presence of hemin, moreover, the terminal sequence of the formed complex could assemble into hemin/G-quadruplex. Therefore, the highly efficient ECL quenching was achieved due to the multiple quenching mechanisms, including electron/energy transfer between ferrocene and QDs, the steric hindrance effects, and the horseradish peroxidase-mimicking activity of hemin/G-quadruplex. Furthermore, owing to the flexibility in regulating the recognition sequences of MNAzyme, the assaying targets can be programmed. Based on the cascade amplification and multiple ECL quenching mechanisms, the developed programmable "signal-off" ECL sensing platform demonstrates excellent sensitivity and the detection limits of 35.00 aM, 3.71 fM, and 0.28 pM (S/N = 3) for target DNA, aptamer substrate (ATP as a model), and ion (Ag+ as a model), respectively.

Holey nitrogen-doped graphene aerogel for simultaneously electrochemical determination of ascorbic acid, dopamine and uric acid.

In this paper, holey nitrogen-doped graphene aerogel (HNGA) was synthesized and applied to the concurrently electrochemical determination of small biological molecules including ascorbic acid (AA), dopamine (DA) and uric acid (UA). Firstly, holey graphene hydrogel was synthesized by the hydrothermal reaction in the presence of H2O2, which subsequently was lyophilized and further annealed in the mixed gas of ammonia and argon to obtain HNGA. Electron microscopy characterization exhibited a great number of nanopores formed on the basal surface of graphene sheets, and HNGA possessed a hierarchically porous structure. The unique structure and composition of HNGA make it an ideal material for electroanalytical application through accelerating mass and electron transfer. HNGA modified glassy carbon electrode (HNGA/GCE) displayed significantly enhanced electrochemical response to AA, DA, and UA, namely reducing overpotential, increasing current density, and improving the reversibility. The oxidation peaks of these three biomolecules can be entirely separated with evident peak potential differences which are 0.216 V (AA-DA), 0.120 V (DA-UA), and 0.336 V (AA-UA), which it allowed the determination of the three substances at the same time. This sensor shows high sensitivity for the determination of AA, DA, and UA with the detection limit of 16.7 µM, 0.22 µM, and 0.12 µM (S/N = 3), respectively. The proposed sensor was applicable for the practical sample analysis as well and desirable recovery was obtained.

The endothelial dysfunction in patients with type 2 diabetes mellitus is associated with IL-6 gene promoter polymorphism in Chinese population.

The purpose of this study is to examine the effects of IL-6 gene promoter -174G/C and -572G/C polymorphism on endothelial function of Chinese T2DM and normal glucose regulation (NGR) subjects. 512 newly diagnosed T2DM patients and 483 NGR subjects were recruited and Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) was performed for the IL-6 gene promoter -174G/C and -572G/C polymorphism. Flow-mediated dilation (FMD) was measured as a non-invasive indicator for endothelial function. The results show that the C allele and CC genotype at -174 of IL-6 gene promoter region was extremely rare in both T2DM and NGR groups; genotypes' and alleles' frequency at -572 of IL-6 gene promoter region is of no difference between T2DM and NGR groups; within T2DM group, higher plasma IL-6 concentration and lower FMD was found in patients with -572 GC (2.36 ± 0.69, 4.23 ± 3.82%) or GG (2.32 ± 0.74, 4.24 ± 3.67%) genotype, compared with patients with CC (2.15 ± 0.62, 5.28 ± 3.94%) genotype. The conclusion of the study is that in comparison with patients of CC genotype, the T2DM patients of -572 GC or GG genotype may have more aggravated endothelial dysfunction (ED) and be at higher risk for coronary artery disease (CAD).