Zhixue Zhou

A new approach to light up DNA/Ag nanocluster-based beacons for bioanalysis

Dynamic DNA assembly, operated in an autonomous and reconfigurable manner by controlling the kinetics of strand displacement reactions (SDR), is an ideal approach to amplify the fluorescent signals for molecular diagnostic and imaging. Herein, we for the first time have demonstrated an enhancement of fluorescence intensity of DNA/Ag nanocluster-based beacons by the modulation of SDR. This is a new DNA/Ag NCs fluorescence light-up system through the use of the enhancer of G-rich overhang. Such a sensing system can be used to develop a DNA/Ag nanocluster-based beacon for the fluorescent detection of nucleic acid and thrombin with high selectivity and sensitivity, in which the detection sensitivity could be further enhanced through additional Exo III based amplification.

Label-free G-quadruplex-specific fluorescent probe for sensitive detection of copper(II) ion.

An effective G-quadruplex-based probe has been constructed for rapid and sensitive detection of Cu(2+). In this probe, an anionic porphyrin, protoporphyrin IX (PPIX) served as a reference signal, which binds to G-quadruplex specifically and the fluorescence intensity increases sharply. While, in the presence of Cu(2+), the G-quadruplex can catalyze the related Cu(2+) insertion into the protoporphyrin, the fluorescent intensity is decreased. The fluorescence of the response ligand could be selectively quenched in the presence of Cu(2+) and not interfered by other metal ions. The probe provided an effective platform for reliable detection of Cu(2+) with a detection limit as low as 3.0nM, the high sensitivity was attributed to the strong metalation of PPIX with Cu(2+) catalyzed by G-quadruplex (PS5.M). Linear correlations were obtained over the logarithm of copper ion concentration in the range from 8×10(-9)M to 2×10(-6)M (R=0.998). The G-quadruplex-based probe also could be used to detect Cu(2+) in real water samples. Additionally, these striking properties endow the G-quadruplex-ligand with a great promise for analytical applications.

Aptamer-Based Sensing Platform Using Three-Way DNA Junction-Driven Strand Displacement and Its Application in DNA Logic Circuit

We proposed a new three-way DNA junction-driven strand displacement mode and fabricated an aptamer-based label-free fluorescent sensing platform on the basis of this mechanism. Assembling the aptamer sequence into the three-way DNA junction makes the platform sensitive to the target of the aptamer. A label-free signal readout method, split G-quadruplex enhanced fluorescence of protoporphyrin IX (PPIX), was used to report the final signal. Here, adenosine triphosphatase (ATP) was taken as a model and detected through this approach, and DNA strand could also be detected by it. The mechanism was investigated by native polyacrylamide gel electrophoresis. Furthermore, on the basis of this molecular platform, we built a logic circuit with ATP and DNA strands as input. Aptamer played an important role in mediating the small molecule ATP to tune the DNA logic gate. Through altering the aptamer sequence, this molecular platform will be sensitive to various stimuli and applied in a wide field.

Label-free aptamer biosensor for thrombin detection based on functionalized graphene nanocomposites

A label-free and amplified electrochemical impedimetric aptasensor based on functionalized graphene nanocomposites (rGO-AuNPs) was developed for the detection of thrombin, which played a vital role in thrombosis and hemostasis. The thiolated aptamer and dithiothreitol (TBA15-DTT) were firstly immobilized on the gold electrode to capture the thrombin molecules, and then aptamer functionalized graphene nanocomposites (rGO-TBA29) were used to fabricate a sandwich sensing platform for amplifying the impedimetric signals. As numerous negative charges of TBA29 on the electrode repelled to the [Fe(CN)6](4-/3-) anions, resulting in an obvious amplified charge-transfer resistance (Rct) signal. The Rct increase was linearly proportional to the thrombin concentration from 0.3 to 50nM and a detection limit of 0.01nM thrombin was achieved. In addition, graphene could also be labeled with other probes via electrostatic or π-π stacking interactions to produce signals, therefore different detection methods expanding wide application could be used in this model.

Paper-based solid-state electrochemiluminescence sensor using poly(sodium 4-styrenesulfonate) functionalized graphene/nafion composite film.

Herein, highly efficient solid-state ECL sensor was introduced for the first time onto the screen printed electrodes of the paper-based chips (PCs) based on the composite film of poly(sodium 4-styrenesulfonate) functionalized graphene (PSSG) and Nafion. Attributed to the cooperative characteristics of both PSS and graphene, PSSG ensured both effective Ru(bpy)(3)(2+) immobilization and fast electron transfer of Ru(bpy)(3)(2+) in the composite film. The ECL behaviors at the developed sensor were investigated using tripropylamine as a representative analyte and low detection limit (SN(-1)=3) of 5.0 nM was obtained. It also exhibited more excellent reproducibility (relative standard deviations of 0.63% for continuous 45 cycles) and long-term stability (~80% of its initial ECL intensity could be retained over 3 months). More importantly, assisted by the developed ECL sensor, discrimination of 1.0 nM single-nucleotide mismatch in human urine matrix could be realized on the PCs for the first attempt. Thus, the developed sensor was confirmed with the advantages of highly sensitivity, long-term stability, simplicity, low cost, disposability, high efficiency and potential applicability.

Molecular aptamer beacon tuned DNA strand displacement to transform small molecules into DNA logic outputs

A molecular aptamer beacon tuned DNA strand displacement reaction was introduced in this work. This strand displacement mode can be used to transform the adenosine triphosphate (ATP) input into a DNA strand output signal for the downstream gates to process. A simple logic circuit was built on the basis of this mechanism.

Engineering DNA Three-Way Junction with Multifunctional Moieties: Sensing Platform for Bioanalysis

Functionalization of DNA nanostructures is critical to the achievement of the application in biosensors. Herein, we demonstrate a novel DNA three-way junction (TWJ) with three functional moieties, which integrates the electrochemical, fluorescent, and colorimetric properties. Upon addition of external stimuli, including DNA, thrombin, and ATP, the specific interactions between targets and sensing elements could induce strand displacement reaction and conformation transformation, resulting in the integration of G-quadruplex/hemin complex as electrochemical probe, lighting up the fluorescence of DNA/Ag nanoclusters and enhancing the catalytic activity of DNAzyme to catalyze the H2O2-TMB system to generate colorimetric signal. Such a functional DNA nanostructure actually serves as a biosensing platform, showing high sensitivity and selectivity for DNA, thrombin, and ATP detection. Furthermore, We also show that this novel sensing platform can be utilized to detect three different kinds of targets independently and simultaneously. Our integrated concept provides a paradigm for exploring the potential of TWJs in analytical applications.

A novel electrochemical sensing platform for anions based on conducting polymer film modified electrodes integrated on paper-based chips.

In this paper, conducting polymer film modified electrodes were applied to fabricate paper-based chips (PCs), and different concentrations of chloride ions (Cl(-)) in water can be selectively detected based on the potential response towards Cl(-). The three-electrode system was screen-printed on paper and the polypyrrole (PPy) film doped with Cl(-) was electrochemically polymerized on working electrodes through cyclic voltammetry in aqueous solution. Open circuit potential-time method was used to measure the potential response. Based on such PCs, Cl(-) can be selectively detected in the range of 10(-7)-10(-2)M. Moreover, such PCs were utilized for Cl(-) analysis in real water samples and resulted in good results with recoveries between 113% and 124%. Besides, following the strategy we also employed this method to detect F(-) in water to demonstrate its general applicability. In view of its novelty, simplicity, sensitivity and low price, such PCs will potentially be utilized for the monitoring of anions in the environment, and our method made a start for the application of CMEs to PCs to design electrochemical sensors.

G-quadruplex DNA/protoporphyrin IX-based synergistic platform for targeted photodynamic cancer therapy

Photodynamic therapy (PDT) is an emerging technique to induce cancer cell death. However, the tumor specificity, cellular uptake and biodistribution of many photosensitizers urgently need to be improved. In this regard, we show here that the integrated nanoassemblies based on G-quadruplex DNAs (GQDs)/protoporphyrin IX (PPIX) can serve as a synergistic platform for targeted high-performance PDT. In the nanoassemblies, GQDs function as carriers of sensitiser PPIX and confers the system cancer cell targeting ability. After nucleolin-mediated efficient binding and cellular uptake of GQDs/PPIX assemblies, the strong red fluorescence of GQDs/PPIX complex provides a powerful tool for biological imaging. Moreover, the reactive oxygen species (ROS) generated by GQDs/PPIX under light illumination can effectively kill cancer cells. The present approach is simply composed by DNA and photosensitizers, thereby avoiding any complicated and time-consuming covalent modification or chemical labeling procedure.