Lingfeng Gao

Dynamically Tunable Chemiluminescence of Luminol-Functionalized Silver Nanoparticles and Its Application to Protein Sensing Arrays

It is still a great challenge to develop an array-based sensing system that can obtain only multiparameters, according to a single experiment and device. The role of conventional chemiluminescence (CL) in biosensing has been limited to a signal transducer in which a single signal (CL intensity) can be obtained for quantifying the concentrations of analytes. In this work, we have developed an dynamically tunable CL system, based on the reaction of luminol-functionalized silver nanoparticles (luminol-AgNPs) with H2O2, which could be tunable via adjusting various conditions such as the concentration of H2O2, pH value, and addition of protein. A single experiment operation could obtain multiparameters including CL intensity, the time to appear CL emission and the time to reach CL peak value. The tunable, low-background, and highly reproducible CL system based on luminol-AgNPs is applied, for the first time, as a sensing platform with trichannel properties for protein sensing arrays by principal component analysis. Identification of 35 unknowns demonstrated a success rate of >96%. The developed sensing arrays based on the luminol-AgNPs provide a new way to use nanoparticles-based CL for the fabrication of sensing arrays and hold great promise for biomedical application in the future.

A general chemiluminescence strategy for measuring aptamer-target binding and target concentration.

Although much effort has been made for studies on aptamer-target interactions due to promising applications of aptamers in biomedical and analytical fields, measurement of the aptamer-target binding constant and binding site still remains challenging. Herein, we report a sensitive label-free chemiluminescence (CL) strategy to determine the target concentration and, more importantly, to measure the target-aptamer binding constant and binding site. This approach is suitable for multiple types of targets, including small molecules, peptides, and proteins that can enhance the CL initiated by N-(aminobutyl)-N-ethylisoluminol functionalized gold colloids, making the present method a general platform to investigate aptamer-target interactions. This approach can achieve extremely high sensitivity with nanogram samples for measuring the target-aptamer binding constant. And the measurement could be rapidly performed using a simple and low-cost CL system. It provides an effective tool for studying the binding of biologically important molecules to nucleic acids and the selection of aptamers. Besides, we have also discovered that the 14-mer aptamer fragment itself split from the ATP-binding aptamer could selectively capture ATP. The binding constant, site, and conformation between ATP and the 14-mer aptamer fragment were obtained using such a novel CL strategy and molecular dynamic simulation.

Highly Chemiluminescent Graphene Oxide Hybrids Bifunctionalized by N-(Aminobutyl)-N-(Ethylisoluminol)/Horseradish Peroxidase and Sensitive Sensing of Hydrogen Peroxide

N-aminobutyl-N-ethylisoluminol and horseradish peroxidase bifunctionalized graphene oxide hybrids (ABEI-GO@HRP) were prepared through a facile and green strategy for the first time. The hybrids exhibited excellent chemiluminescence (CL) activity over a wide range of pH from 6.1 to 13.0 when reacted with H2O2, whereas ABEI functionalized GO had no CL emission at neutral pH and showed more than 2 orders of magnitude lower CL intensity than ABEI-GO@HRP at pH 13.0. Such strong CL emission from ABEI-GO@HRP was probably due to that HRP and GO facilitated the formation of O2(•-), - CO4(•2-), HO(•), and π-C═C(•) in the CL reaction, and GO as a reaction interface promoted the electron transfer of the radical-involved reaction. By virtue of ABEI-GO@HRP as a platform, an ultrasensitive, selective, and reagentless CL sensor was developed for H2O2 detection. The CL sensor exhibited a detection limit of 47 fM at physiological pH, which was more than 2 orders of magnitude lower than previously reported methods. This work reveals that bifunctionalization of GO by ABEI and HRP leads to excellent CL feature and enzyme selectivity, which can be used as an ideal platform for developing novel analytical methods.

A general strategy to prepare homogeneous and reagentless GO/lucigenin&enzyme biosensors for detection of small biomolecules.

In this work, a novel biosensor was developed for the detection of glucose based on glucose oxidase (GOD) functionalized graphene oxide (GO)/lucigenin nanocomposite. In this sensing strategy, GO/lucigenin composite was first prepared by vigorously stirring GO with lucigenin. Then the functionalization of GOD was achieved by simply storing GOD with GO/lucigenin at 4 °C overnight to form GO/lucigenin&GOD composite. When glucose was incubated with GO/lucigenin&GOD composite for 50 min to generate H2O2, followed by the injection of 0.2M NaOH, CL signal was detected due to the reaction of lucigenin with H2O2. Glucose could be determined in the range of 1.0×10(-6)-5.0×10(-3) g mL(-1) with a detection limit of 9.9×10(-7) g mL(-1). The present biosensor has been successfully applied for the detection of glucose in human serum samples. Compared with previously reported methods, this sensing strategy is homogeneous and reagentless and avoids complicated assembly procedure and pretreatment of serum sample, showing good stability, repeatability, high selectivity and simplicity. Moreover, this strategy has been demonstrated to be a general strategy by replacing GOD with other enzymes such as uricase and choline oxidase for the detection of small molecules such as uric acid and choline. The proposed biosensors may find future applications in the fields such as disease diagnosis and biomedicine.

A label-free method for the detection of specific DNA sequences using gold nanoparticles bifunctionalized with a chemiluminescent reagent and a catalyst as signal reporters

AbstractSensitive, specific, simple, fast, and low-cost DNA detection methods are extremely important in clinical diagnostics, gene therapy, and a variety of biomedical studies. In this work, we developed a general method for the detection of specific DNA sequences from Mycobacterium tuberculosis (TB), hepatitis B virus (HBV), and myelocytomatosis viral oncogene (v-myc) using gold nanoparticles bifunctionalized with bothxa0a chemiluminescent (CL) reagent and a catalytic metal complex as signal reporters and a DNA strand complementary to the target as the capture probe. In this CL method, a biotinylated single-strand DNA capture probe was immobilized in a streptavidin-coated microwell. Upon the addition of the target single-strand DNA, the capture probe hybridized with the target DNA. After adding the bifunctionalized gold nanoparticles and H2O2, a well-defined CL signal was obtained, and the CL intensity was observed to change as the target DNA concentration was increased. It was possible to determine the concentration of the target TB single-strand DNA in the range 1.0u2009×u200910−13–1.0u2009×u200910−8 M with a detection limit of 4.8u2009×u200910−14 M. HBV single-strand DNA and v-myc single-strand DNA could also be determined in the range 1.0u2009×u200910−11–1.0u2009×u200910−8 M with detection limits of 5.9u2009×u200910−12 M and 8.0u2009×u200910−12 M, respectively, using this CL technique. The method reported in this paper is the first label-free CL method for the determination of specific DNA sequences to utilize gold nanoparticles bifunctionalized with both a CL reagent and a catalytic metal complex. The sensitivity of this CL method is superior to those of most previously reported label-free methods. Compared with methods that use polymerase chain reaction amplification, this label-free CL method is much simpler, faster, and more economic. This work has thus demonstrated a simple and fast scanning strategy for the detection of specific DNA sequences related to diseases.n Graphical AbstractSchematic illustration of label-free CL method for detection of specific DNA sequences

Firefly-mimicking intensive and long-lasting chemiluminescence hydrogels

Most known chemiluminescence (CL) reactions exhibit flash-type light emission. Great efforts have been devoted to the development of CL systems that emit light with high intensity and long-lasting time. However, axa0long-lasting CL system that can last for hundreds of hours is yet-to-be-demonstrated. Here we show firefly-mimicking intensive and long-lasting CL hydrogels consisting of chitosan, CL reagent N-(4-aminobutyl)-N-ethylisoluminol (ABEI) and catalyst Co2+. The light emission is even visible to naked eyes and lasts for over 150u2009h when the hydrogels are mixed with H2O2. This is attributed to slow-diffusion-controlled heterogeneous catalysis. Co2+ located at the skeleton of the hydrogels as an active site catalyzes the decomposition of slowly diffusing H2O2, followed by the reaction with ABEI to generate intensive and long-lasting CL. This mimics firefly bioluminescence system in terms of intensity, duration time and catalytic characteristic, which is of potential applications in cold light sources, bioassays, biosensors and biological imaging.Great efforts have been devoted to the development of chemiluminescence systems that emit light with high intensity over long periods of time. Here the authors show, firefly-mimicking intensive and long-lasting chemiluminescence hydrogels consisting of chitosan, N-(4-aminobutyl)-N-ethylisoluminol (ABEI) and catalyst Co2+.

Chemiluminescent and fluorescent dual-signal graphene quantum dots and their application in pesticide sensing arrays

In this work, N-(aminobutyl)-N-(ethylisoluminol) (ABEI) functionalized graphene quantum dots (ABEI-GQDs) with excellent chemiluminescence (CL) and fluorescence (FL) properties were synthesized. Firstly, small 2-dimensional graphene oxide (GO) was prepared by exfoliating and crushing graphite oxide in the presence of HNO3 and KMnO4. Then the size of small 2-dimensional GO was further decreased by reduction with NaBH4 to form GQDs. After that, ABEI was immobilized to the surface of GQDs by simply stirring the mixture of GQDs and ABEI. The synthesized ABEI-GQDs demonstrated excellent CL properties when reacting with H2O2 and wavelength-tunable FL emission with increasing excitation wavelength. And the bright blue FL of ABEI-GQDs could be seen even with the naked eye. Based on their CL and FL properties, a pesticide-sensing array was developed for the differentiation of pesticides. It was found that the sensing strategy could distinguish five pesticides with different concentrations, including thiamethoxam, flubendiamide, dimethoate, dipterex and chlorpyrifos. Novel CL and FL dual-signal graphene quantum dots ABEI-GQDs were obtained for the first time, which may find more applications in bioassays and bioimaging in the future.

Acridinium Ester-Functionalized Carbon Nanomaterials: General Synthesis Strategy and Outstanding Chemiluminescence

In this work, three different kinds of acridinium ester (AE)-functionalized carbon nanomaterials, including AE-functionalized carbon nanoparticles (AE-CNPs), AE-functionalized graphene oxide (AE-GO), and AE-functionalized multiwalled carbon nanotubes (AE-MCNTs), were synthesized for the first time via a simple, general, and noncovalent strategy. AE molecules were assembled on the surface of carbon nanomaterials by electrostatic interaction, π-π stacking interaction, and amide bond. The synthesized AE-CNPs, AE-GO, and AE-MCNTs with 5.0 × 10(-8) mol·L(-1) of synthetic AE concentration, which was very low compared with other chemiluminescence (CL) reagents such as luminol, N-(aminobutyl)-N-(ethylisoluminol), and lucigenin at the concentration of 3.3 × 10(-4) to 5.0 × 10(-6) mol·L(-1) used for the synthesis of CL-functionalized nanomaterials, exhibited outstanding CL activity and good stability. It was found that carbon nanomaterials as nanosized platforms could efficiently immobilize AE molecules and facilitate the formation of OH(•) and O2(•-), leading to strong light emission. Moreover, the CL intensity of AE-GO was the highest, which was about 8.7 and 3.7 times higher than that of AE-CNPs and AE-MCNTs, respectively. This mainly resulted from a difference in the amount of adsorbed AE molecules on the surface of different carbon nanomaterials. Additionally, the prepared AE-CNPs demonstrated excitation-dependent fluorescence property and good fluorescence stability against photobleaching. On the basis of the excellent CL and special fluorescence properties of AE-CNPs, a dual-mode array strategy has been proposed for the first time and seven kinds of transition-metal ions could be successfully discriminated.

Highly Chemiluminescent Magnetic Beads for Label-Free Sensing of 2,4,6-Trinitrotoluene

Until now, despite the great success acquired in scientific research and commercial applications, magnetic beads (MBs) have been used for nothing more than a carrier in most cases in bioassays. In this work, highly chemiluminescent magnetic beads containing N-(4-aminobutyl)-N-ethyl isoluminol (ABEI) and Co2+ (Co2+/ABEI/MBs) were first synthesized via a facile strategy. ABEI and Co2+ were grafted onto the surface of carboxylated MBs by virtue of a carboxyl group and electrostatic interaction. The as-prepared Co2+/ABEI/MBs exhibited good paramagnetic properties, satisfactory stability, and intense chemiluminescence (CL) emission when reacted with H2O2, which was more than 150 times that of ABEI functionalized MBs. Furthermore, it was found that 2,4,6-trinitrotoluene (TNT) aptamer could attach to the surface of Co2+/ABEI/MBs via electrostatic interaction and coordination interaction between TNT aptamer and Co2+, leading to a decrease in CL intensity due to the catalytic site Co2+ being blocked by the aptamer. In the presence of TNT, TNT would bind strongly with TNT aptamer and detach from the surface of Co2+/ABEI/MBs, resulting in partial restoration of the CL signal. Accordingly, label-free aptasensor was developed for the determination of TNT in the range of 0.05-25 ng/mL with a detection limit of 17 pg/mL. This work demonstrates that Co2+/ABEI/MBs are easily connected with recognition biomolecules, which are not only magnetic carriers but also direct sensing interfaces with excellent CL activity. It provides a novel CL interface with a magnetic property which easily separates analytes from the sample matrix to construct label-free bioassays.

Determination of the binding constant of specific interactions and binding target concentration simultaneously using a general chemiluminescence method

The measurement of the binding constant of specific interactions and concentration of a target is of considerable importance in clinical diagnosis, therapy, bioassays and drug design. The development of methods that combine high sensitivity with generalization and simplicity for the measurement of both binding constant and target concentration is highly desirable. Previously, we developed a label-free chemiluminescence (CL) strategy for the measurement of the target concentration and binding constants between DNA aptamers and target simultaneously based on the fact that the target could enhance the CL produced by the reaction of the N-(4-aminobutyl)-N-ethylisoluminol (ABEI) functionalized gold colloid with H2O2. In this study, the enhancement and inhibition effect of various targets on the CL reaction is studied. The generalization of the proposed CL strategy for various targets is also explored. The results demonstrate that the proposed CL strategy is suitable for targets that can cause a change in CL intensity, which includes enhancement and inhibition. It could be applied for the measurement of the dissociation constants of aptamer-binding targets, antibody–antigen complexes, protein-binding small molecules and double-strand DNA hybrids from the millimole to picomole level. It could also be used for the sensitive determination of target concentration, including 2,4,6-trinitrotoluene (TNT), dopamine, tetracycline, human IgG (hIgG), tuberculosis (TB) DNA and mannose, with the detection limit of 0.93 nM–4.1 fM. This strategy is of great potential in fundamental research as well as in applications in life sciences.