Co-reporter:Wenshan Li;Lei Luo;Jin Huang;Qing Wang;Jianbo Liu;Xiao Xiao;Hongmei Fang;Kemin Wang
Chemical Communications 2017 vol. 53(Issue 40) pp:5565-5568
Publication Date(Web):2017/05/17
DOI:10.1039/C7CC01128H
We report self-assembled nanocentipedes as multivalent vehicles for the delivery of immunostimulatory CpG ODNs. The multivalent vehicles could be internalized by RAW264.7 cells and stimulate the secretion of large amounts of cytokines, successively inducing effective apoptosis of cancer cells.
Co-reporter:Chunxia Song;Bingjie Li;Kemin Wang;Qing Wang;Jianbo Liu;Jin Huang
Analyst (1876-Present) 2017 vol. 142(Issue 1) pp:224-228
Publication Date(Web):2016/12/19
DOI:10.1039/C6AN02269C
Herein, we proposed an enzyme-free strategy for the amplified detection of DNA by combining the efficient fluorescence enhancement capability of a β-cyclodextrin-tethered cationic polymer (cationic polyβ-CD) to pyrene with the amplification capability of target DNA triggered hybridization chain reaction (HCR). Cationic polyβ-CD with positive charge was synthesized. Two hairpin probes, H1 and H2, were employed in the system and the pyrene-labelled H2 was chosen as the signal unit. The pyrene attached on the sticky end of H2 was flexible and there was strong electrostatic interaction between cationic polyβ-CD and negatively-charged H2, so pyrene could easily enter the cavity of CD that is tethered on the cationic polymer, accompanied by significant fluorescence enhancement. Once target DNA was introduced, HCR was triggered to form a rigid long dsDNA polymer with pyrene attached on it. The pyrene was hardly able to enter the cavity of cationic polyβ-CD because of steric hindrance, leading to a weak fluorescent signal. Owing to the efficient pyrene fluorescence enhancement of cationic polyβ-CD and the amplified capability of HCR, an enzyme-free sensitive detection of target DNA was achieved with a detection limit of 0.1 nM and high selectivity.
Co-reporter:Shiya Qin, Nandi Chen, Xiaohai Yang, Qing Wang, Kemin WangJin Huang, Jianbo Liu, Maogui Zhou
ACS Sensors - New in 2016 2017 Volume 2(Issue 2) pp:
Publication Date(Web):January 20, 2017
DOI:10.1021/acssensors.6b00836
Pancreatic polypeptide (PP) is a specific biomarker of nonfunctional pancreatic neuroendocrine tumors (NF-pNETs). Clinical significance of PP inspires researchers to make great efforts in developing sensitive and specific sensors. However, there is no existing biosensor for detecting PP that combines facility and functionality. Addressing this challenge, a pair of aptamers which could be used to develop a sandwich assay for PP is reported. First, several high affinity aptamers are screened through graphene oxide-based SELEX, and appropriate dual-aptamers which could bind to different epitopes of PP are identified through fluorescence assays. Then the feasibility of the dual-aptamers for constructing the sandwich assay is validated via dynamic light scattering. This sandwich assay shows considerable sensitivity and specificity. The above results imply that the dual-aptamers have the potential toward developing novel sensors for PP in clinical samples.Keywords: aptamer; gold nanoparticle; graphene oxide; pancreatic polypeptide; sandwich assay;
Co-reporter:Wenshan Li, Xiaohai Yang, Leiliang He, Kemin Wang, Qing Wang, Jin Huang, Jianbo Liu, Bin Wu, and Congcong Xu
ACS Applied Materials & Interfaces 2016 Volume 8(Issue 39) pp:25733
Publication Date(Web):September 13, 2016
DOI:10.1021/acsami.6b08210
An idea drug carrier, with good binding affinity, selectivity, drug payload capacity, and cellular internalized capability, will greatly improve the efficiency of target delivery. Herein a self-assembled and multivalent DNA nanostructure was developed as drug carrier for efficient and targeted delivery. The DNA structure was similar to that of a centipede, composed of trunk and legs: The trunk was a self-assembled DNA scaffold via hybridization chain reaction (HCR) from two biotinylated hairpin monomers created upon initiation by a trigger DNA, and the legs were biotinylated aptamers conjugated to the trunk via streptavidin–biotin affinity interaction. The long trunk of the “DNA nanocentipede” was loaded with doxorubicin (Dox), and the legs were SMMC-7721 cell-binding aptamers (Zy1) which functioned as targeting moieties to firmly and selectively grasp target cells. The results of agarose gel electrophoresis and fluorescence anisotropy confirmed that Zy1-based DNA nanocentipedes (Zy1-Nces) were successfully constructed. Flow cytometric analyses demonstrated that Zy1-Nces were more effective than free Zy1 in binding affinity and selectivity due to a multivalent effect. Confocal microscopy studies demonstrated that the internalization was highly dependent on the higher valences of DNA nanocentipedes without the loss of selectivity. Meanwhile, Zy1-Nces exhibited high drug-loading capacity and selective drug transport. The results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed enhanced cellular cytotoxicity of the Dox-loaded Zy1-Nces (Zy1-Nces-Dox) to the target SMMC-7721 cells but not negative control L02 cells. This approach is applicable to prepare drug carriers for other targets by construction of the nanocentipedes with relevant nucleic acid fragments.Keywords: aptamer; cancer theranostics; multivalent; self-assembly; targeted delivery
Co-reporter:Nandi Chen, Shiya Qin, Xiaohai Yang, Qing Wang, Jin Huang, and Kemin Wang
ACS Applied Materials & Interfaces 2016 Volume 8(Issue 40) pp:26552
Publication Date(Web):September 21, 2016
DOI:10.1021/acsami.6b08695
DNA nanostrucures are promising materials for biomedical applications. Herein, we established a “sense-and-treat” localized drug delivery system based on a DNA nanodevice to specifically destroy circulating tumor cells (CTCs) by synergetic chemotherapy and photodynamic therapy. The DNA nanodevices could sense the existence of CTCs and treat CTCs with anticancer agents. Typically, the presence of target cell promoted the formation of hairpin structure of aptamer, and then the aptamer-accompanied DNA tetrahedron would release from the supporter. The chemotherapy drugs (doxorubicin, Dox) loaded in DNA tetrahedron would destroy the CTCs specifically. Moreover, the photosensitizer labeled on DNA tetrahedron would be activated by lights and generated toxic 1O2, once DNA nanodevices bound CTCs flow through the superficial capillary. Unlike the aptamer only labeled with photosensitizer, the DNA nanodevice showed the capability to promote cellular internalization of anticancer agents, increase drug loading capacity, and realize synergetic therapy, which enhanced the destructive ability of anticancer agents. As proof of concept, this DNA nanodevice has the potential to inhibit metastasis by synergetic destruction of CTCs.Keywords: aptamer; chemotherapy; circulating tumor cell; DNA nanodevice; metastasis; photodynamic therapy
Co-reporter:Nandi Chen, Xiaohai Yang, Qing Wang, Lixin Jian, Hui Shi, Shiya Qin, Kemin Wang, Jin Huang and Wenjing Liu
Chemical Communications 2016 vol. 52(Issue 41) pp:6789-6792
Publication Date(Web):20 Apr 2016
DOI:10.1039/C6CC02374F
Existing drug delivery systems were not suitable for killing cells in the circulatory system specifically. Herein, we developed a novel localized drug delivery strategy, in which the release of anticancer agents was specifically triggered by circulating tumor cells. Meanwhile, damage to non-target cells was avoided.
Co-reporter:Bin Wu, Rui Jiang, Qing Wang, Jin Huang, Xiaohai Yang, Kemin Wang, Wenshan Li, Nandi Chen and Qing Li
Chemical Communications 2016 vol. 52(Issue 17) pp:3568-3571
Publication Date(Web):27 Jan 2016
DOI:10.1039/C5CC10486F
High affinity DNA aptamers against C-reactive protein (CRP) were obtained using a microfluidic chip. The aptamers were then used for the construction of an Au nanoparticle enhanced surface plasmon resonance biosensor, which was introduced for the detection of CRP at concentrations ranging from 10 pM to 100 nM in diluted human serum.
Co-reporter:Haihua Huang, Xiaohai Yang, Kemin Wang, Qing Wang, Qiuping Guo, Jin Huang, Jianbo Liu and Chunxia Song
Analyst 2016 vol. 141(Issue 8) pp:2502-2507
Publication Date(Web):07 Mar 2016
DOI:10.1039/C5AN02658J
Nowadays, enzyme-free nucleic acid-based signal amplification strategies are frequently utilized in the design of biosensors due to their excellent sensitivity. Developing more extended analytical methods is fundamental for basic studies in the biological and biomedical fields. Herein, we introduce an enzyme-free amplified detection strategy for the small molecule adenosine. The approach is based on adenosine–aptamer binding triggered catalyzed hairpin assembly and host–guest interactions between β-cyclodextrin polymer (β-CDP) and pyrene. Two hairpin probes (probe H1 and probe H2) and an aptamer-trigger/inhibitor duplex probe were employed in the system and the pyrene-labeled probe H1 was chosen as the signal unit. In the absence of adenosine, the aptamer-trigger was inhibited by the inhibitor strand. The hairpin probes were in the closed hairpin formation without activation of the trigger strand. Pyrene labeled at the 5′-termini of the single-stranded stem of probe H1 could be easily trapped in the hydrophobic cavity of β-CDP because of weak steric hindrance, leading to a significant fluorescence enhancement. Once the hairpin assembly was catalyzed by the adenosine–aptamer binding event, a hybridized DNA duplex H1/H2 was created continuously. Pyrene labeled at the 5′-termini of the DNA duplex H1/H2 finds it difficult to enter the cavity of β-CDP due to steric hindrance, leading to a weaker fluorescence signal. Thus, the target could be detected by this simple mix-and-detect amplification method without a need for expensive and perishable protein enzymes. As low as 42 nM of adenosine was detected by this assay, which is comparable to that of some reported colorimetric methods. Meanwhile, the proposed method was further successfully applied to detect adenosine in human serum samples, showing great potential for adenosine detection from complex fluids.
Co-reporter:Wenjing Liu, Min Lin, Xiaohai Yang, Bin Wu, Nandi Chen, Qing Wang, Kemin Wang and Shiya Qin
Analyst 2016 vol. 141(Issue 14) pp:4463-4469
Publication Date(Web):13 May 2016
DOI:10.1039/C6AN00987E
G-quadruplexes are guanine-rich nucleic acid sequences that can act as universal signal-transducers and generate colorimetric, fluorescence, and chemiluminescence signals when complexed with different ligands. Due to their merits including easy modification and low cost, it is of great importance to explore new G-quadruplexes with improved performance. Herein the properties of newly identified G-quadruplexes 9th-3-35 and 10th-2-40 were investigated in detail with UV-vis spectra, circular dichroism (CD) spectra and fluorescence spectra. The results indicated that 9th-3-35 and 10th-2-40 exhibited excellent peroxidase-like activity, as well as fluorescence enhancement of thioflavin T (ThT). Furthermore, the application of G-quadruplexes to DNA detection was performed on account of the ThT fluorescence enhancement, and the limit of detection was as low as 8 pM. This study implied that 9th-3-35 and 10th-2-40 are competitive candidates as signal-transducers in the design of bioassays.
Co-reporter:Qing Wang;Bianxia Luo;Kemin Wang;Lin Liu;Shasha Du ;Zhiping Li
Journal of Molecular Recognition 2016 Volume 29( Issue 4) pp:151-158
Publication Date(Web):
DOI:10.1002/jmr.2514
The immobilization strategy of cell-specific aptamers is of great importance for studying the interaction between a cell and its aptamer. However, because of the difficulty of studying living cell, there have not been any systematic reports about the effect of immobilization strategies on the binding ability of an immobilized aptamer to its target cell. Because atomic force spectroscopy (AFM) could not only be suitable for the investigation of living cell under physiological conditions but also obtains information reflecting the intrinsic properties of individuals, the effect of immobilization strategies on the interaction of aptamer/human hepatocarcinoma cell Bel-7404 was successively evaluated using AFM here. Two different immobilization methods, including polyethylene glycol immobilization method and glutaraldehyde immobilization method were used, and the factors, such as aptamer orientation, oligodeoxythymidine spacers and dodecyl spacers, were investigated. Binding events measured by AFM showed that a similar unbinding force was obtained regardless of the change of the aptamer orientation, the immobilization method, and spacers, implying that the biophysical characteristics of the aptamer at the molecular level remain undisturbed. However, it showed that the immobilization orientation, immobilization method, and spacers could alter the binding probability of aptamer/Bel-7404 cell. Presumably, these factors may affect the accessibility of the aptamer toward its target cell. These results may provide valuable information for aptamer sensor platforms including ultrasensitive biosensor design. Copyright © 2015 John Wiley & Sons, Ltd.
Co-reporter:Bin Wu, Nandi Chen, Qing Wang, Xiaohai Yang, Kemin Wang, Wenshan Li, Qing Li, Wenjing Liu and Hongmei Fang
Analytical Methods 2016 vol. 8(Issue 21) pp:4177-4180
Publication Date(Web):03 May 2016
DOI:10.1039/C6AY01007E
We developed a simple label-free aptamer-based method for C-reactive protein (CRP) detection. In our previous work, we obtained a G-rich aptamer named 6th-62-40, which exhibited good affinity and specificity toward CRP. Herein, we demonstrated that 6th-62-40 could enhance the fluorescence of thioflavin T (ThT). In the absence of CRP, ThT bound to the G-rich aptamer and formed an aptamer/ThT complex, which showed high fluorescence. However, the fluorescence decreased due to the formation of an aptamer/CRP complex in the presence of CRP. The proposed method was a rapid strategy for CRP detection with a detection limit of 380 pM.
Co-reporter:Chunxia Song, Xiaohai Yang, Kemin Wang, Qing Wang, Jianbo Liu, Jin Huang, Maogui Zhou, Xiaochen Guo
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 2016 Volume 156() pp:131-137
Publication Date(Web):5 March 2016
DOI:10.1016/j.saa.2015.12.001
•Sensitive•Single-labeling of DNA probe•Potential for screening of inhibitorsWe herein report a strategy for sensitive alkaline phosphatase (ALP) fluorescent sensing based on steric hindrance regulated supramolecular assembly between β-cyclodextrin polymer (polyβ-CD) and pyrene. The fluorescence of pyrene was enhanced more than 10 times through supramolecular assembly with polyβ-CD. The 5′-phosphorylated dsDNA probe with pyrene attached on the 3′-terminal could be cleaved by λ exonuclease (λ exo), yielding pyrene attached on mononucleotides. Pyrene attached on mononucleotides could easily enter the cavity of polyβ-CD, resulting in fluorescence enhancement. When ALP was introduced, it could remove 5′-phosphate groups from dsDNA and then prevented the cleavage of dsDNA. Pyrene attached on dsDNA was difficult to enter the cavity of polyβ-CD because of steric hindrance, resulting in an inconspicuous fluorescence enhancement. Owing to the excellent fluorescence enhancement during steric hindrance regulated supramolecular assembly, excellent performance of the assay method was achieved for ALP with a detection limit of 0.04 U mL− 1. The detection limit was superior or comparable with the reported methods. Besides, this method was simple in design, avoiding double-labeling of probe.
Co-reporter:Qing Wang, Rongjuan Liu, Xiaohai Yang, Kemin Wang, Jinqing Zhu, Leiliang He, Qing Li
Sensors and Actuators B: Chemical 2016 Volume 223() pp:613-620
Publication Date(Web):February 2016
DOI:10.1016/j.snb.2015.09.152
•A novel enzyme-free amplified SPR biosensor for microRNA detection was introduced.•This assay showed excellent sensitivity due to the dual signal amplification.•This approach showed good selectivity, resulting in its application in serum sample.A novel enzyme-free amplified surface plasmon resonance (SPR) biosensor for microRNA (miRNA) detection was developed based on gold nanoparticles (AuNPs) coupled with DNA supersandwich. In the detection strategy, the DNA-linked AuNPs as the primary amplification element, not only hybridized with the capture DNA on the Au film to amplify SPR signal but also initiated the subsequent secondary amplification, i.e. DNA supersandwich formation of two report probes. In the presence of target, stem–loop structure of capture DNA on the Au film surface was unfolded, and DNA-linked AuNPs were bound to Au film by hybridization with terminus of capture DNA. Then, the carried assistant DNA on the AuNPs could trigger an alternative hybridization reaction of two report probes, resulting in the formation of DNA supersandwich. Due to the electronic coupling between localized plasmon of AuNPs and the surface plasmon wave associated with Au film, as well as the enhancement of the refractive index of the medium next to the metal film caused by DNA supersandwich structure, the shift of resonance angle was enhanced obviously. By employing the enzyme-free dual signal amplification strategies, as low as ca. 8 fM miRNA-21 could be detected. Moreover, this assay also showed high selectivity toward single-base mismatch, and demonstrated its applicability for the target detection in human serum. This work may provide great potential applications in future clinical analysis.A novel enzyme-free amplified surface plasmon resonance (SPR) biosensor for microRNA (miRNA) detection was developed based on gold nanoparticles (AuNPs) coupled with DNA supersandwich.
Co-reporter:Min Lin, Wenshan Li, Yaning Wang, Xiaohai Yang, Kemin Wang, Qing Wang, Pei Wang, Yujie Chang and Yuyu Tan
Chemical Communications 2015 vol. 51(Issue 39) pp:8304-8306
Publication Date(Web):09 Apr 2015
DOI:10.1039/C5CC00929D
Discrimination of hemoglobins with subtle differences was achieved using an aptamer based sensing array. Linear discriminant analysis (LDA) showed that the sensing array can discriminate human hemoglobins from hemoglobins of different species.
Co-reporter:Chunxia Song, Xiaohai Yang, Kemin Wang, Qing Wang, Jianbo Liu, Jin Huang, Leiliang He, Pei Liu, Zhihe Qing and Wei Liu
Chemical Communications 2015 vol. 51(Issue 10) pp:1815-1818
Publication Date(Web):10 Dec 2014
DOI:10.1039/C4CC08991J
A strategy for T4 polynucleotide kinase activity detection was proposed based on a β-cyclodextrin polymer (polyβ-CD) and an exonuclease reaction. The fluorescence of pyrene enhanced by more than 10 times in the presence of polyβ-CD, and a simple detection of T4 PNK was achieved with a detection limit of 0.02 units per mL.
Co-reporter:Meng Yang, Xiaohai Yang, Kemin Wang, Qing Wang, Xin Fan, Wei Liu, Xizhen Liu, Jianbo Liu, and Jin Huang
Analytical Chemistry 2015 Volume 87(Issue 3) pp:1544
Publication Date(Web):January 2, 2015
DOI:10.1021/ac503813r
The transport of ionic species through a nanochannel plays important roles in fundamental research and practical applications of the nanofluidic device. Here, we demonstrated that ionic transport selectivity of a positively charged nanochannel membrane can be tuned under a phosphoric acid gradient. When phosphoric acid solution and analyte solution were connected by the positively charged nanochannel membrane, the faster-moving analyte through the positively charged nanochannel membrane was the positively charged dye (methylviologen, MV2+) instead of the negatively charged dye (1,5-naphthalene disulfonate, NDS2–). In other words, a reversed ion selectivity of the nanochannel membranes can be found. It can be explained as a result of the combination of diffusion, induced electroosmosis, and induced electrophoresis. In addition, the influencing factors of transport selectivity, including concentration of phosphoric acid, penetration time, and volume of feed solution, were also investigated. The results showed that the transport selectivity can further be tuned by adjusting these factors. As a method of tuning ionic transport selectivity by establishing phosphoric acid gradient, it will be conducive to improving the separation of ionic species.
Co-reporter:Pei Liu, Shan Sun, Xiaochen Guo, Xiaohai Yang, Jin Huang, Kemin Wang, Qing Wang, Jianbo Liu, and Leiliang He
Analytical Chemistry 2015 Volume 87(Issue 5) pp:2665
Publication Date(Web):January 27, 2015
DOI:10.1021/ac503301q
We developed a novel homogeneous fluorescence analysis based on a novel competitive host–guest interaction (CHGI) mechanism between β-cyclodextrin polymer (polyβ CD) and pyrene-labeled probe for biochemical assay. Pyrene labeling with oligonucleotide strands can be recruited and reside in lipophilic cavities of polyβ CD. This altered lipophilic microenvironment provides favored polarity for enhanced quantum efficiencies and extraordinarily increases the luminescence intensity of pyrene. However, with addition of complementary DNA, the pyrene-labeled probe formed double-strand DNA to hinder pyrene from entering the cavities of polyβ CD. The release of pyrene from polyβ CD, which are followed by fluorescence extinguishing, will provide the clear signal turn-off in the presence of target DNA. We also introduced Exodeoxyribonuclease I (Exo I) and Exodeoxyribonuclease III (Exo III) to improve the sensitivity of this system, and the following product of cleavage reaction, pyrene-nucleotide, could more easily host–guest interact with polyβ CD and emit stronger fluorescence than pyrene-labeled probe. In addition, the successful detection of adenosine is also demonstrated by using the similar sensing scheme. Although this scheme might be easily interfered by some biomolecules in the real test sample, it holds promising potential for detecting a broad range of other types of aptamer-binding chemicals and biomolecules.
Co-reporter:Qing Wang, Lin Liu, Xiaohai Yang, Kemin Wang, Nandi Chen, Chenchen Zhou, Bianxia Luo, and Shasha Du
Analytical Chemistry 2015 Volume 87(Issue 4) pp:2242
Publication Date(Web):January 23, 2015
DOI:10.1021/ac503885e
The effects of medicine on the biomolecular interaction have been given increasing attention in biochemistry and affinity-based analytics since the environment in vivo is complex especially for the patients. Herein, myoglobin, a biomarker of acute myocardial infarction, was used as a model, and the medicine effects on the interactions of myoglobin/aptamer and myoglobin/antibody were systematically investigated using atomic force microscopy (AFM) for the first time. The results showed that the average binding force and the binding probability of myoglobin/aptamer almost remained unchanged after myoglobin-modified gold substrate was incubated with promazine, amoxicillin, aspirin, and sodium penicillin, respectively. These parameters were changed for myoglobin/antibody after the myoglobin-modified gold substrate was treated with these medicines. For promazine and amoxicillin, they resulted in the change of binding force distribution of myoglobin/antibody (i.e., from unimodal distribution to bimodal distribution) and the increase of binding probability; for aspirin, it only resulted in the change of the binding force distribution, and for sodium penicillin, it resulted in the increase of the average binding force and the binding probability. These results may be attributed to the different interaction modes and binding sites between myoglobin/aptamer and myoglobin/antibody, the different structures between aptamer and antibody, and the effects of medicines on the conformations of myoglobin. These findings could enrich our understanding of medicine effects on the interactions of aptamer and antibody to their target proteins. Moreover, this work will lay a good foundation for better research and extensive applications of biomolecular interaction, especially in the design of biosensors in complex systems.
Co-reporter:Leiliang He, Xiaohai Yang, Fang Zhao, Kemin Wang, Qing Wang, Jianbo Liu, Jin Huang, Wenshan Li, and Meng Yang
Analytical Chemistry 2015 Volume 87(Issue 4) pp:2459
Publication Date(Web):January 22, 2015
DOI:10.1021/ac504458r
Self-assembly of small building blocks into functional supramolecular nanostructure has opened prospects for the design of novel materials. With this molecular engineering strategy, we have developed self-assembled supramolecular nanoprobes (SSNPs) for ratiometric fluorescence measurement of pH values in cells. The nanoprobes with a diameter of ∼30 nm could be formulated just by mixing pH-sensitive adamantane–fluorescein (Ad-F) and pH-insensitive adamantane–Rhodamine B (Ad-R) with β-cyclodextrin polymer (poly-β-CD) at one time. The nanoprobes with good biocompatibility have been successfully applied to measure intracellular pH in the pH range of 4–8 and estimate pH fluctuations associated with different stimuli in cells. Moreover, we expect that this self-assembled approach is applicable to the construction of nanoprobes for other targets in cells just by replacing the respective indicator dyes with relevant indicators.
Co-reporter:Xiaochen Guo, Pei Liu, Xiaohai Yang, Kemin Wang, Qing Wang, Qiuping Guo, Jin Huang, Jianbo Liu, Chunxia Song and Wenshan Li
Analyst 2015 vol. 140(Issue 6) pp:2016-2022
Publication Date(Web):03 Feb 2015
DOI:10.1039/C4AN02402H
A multiple amplification strategy has been developed for nucleic acid detection based on host–guest interaction between the β-cyclodextrin polymer (β-CDP) and pyrene. Briefly, the detection system consists of three parts: the polymerase and nicking enzyme-assisted isothermal strand displacement amplification (SDA) activated by a target DNA or microRNA; the exonuclease III-aided cyclic enzymatic amplification (CEA); and the fluorescence enhancement effect based on host–guest interaction between β-CDP and pyrene. This strategy showed a good positive linear correlation with target DNA concentrations in the range from 75 fM to 1 pM with a detection limit of 41 fM. Significantly, our amplification platform was further validated and evaluated successfully by assaying miRNA-21 in human serum. The proposed assay has great potential as a nucleic acid quantification method for use in biomedical research, clinical analysis and disease diagnostics.
Co-reporter:Qing Wang, Hui Wang, Xiaohai Yang, Kemin Wang, Rongjuan Liu, Qing Li and Jinqing Ou
Analyst 2015 vol. 140(Issue 4) pp:1161-1165
Publication Date(Web):05 Dec 2014
DOI:10.1039/C4AN02033B
Assays of α-amylase (AMS) activity in serum and urine constitute the important indicator for the diagnosis of acute pancreatitis, mumps, renal disease and abdominal disorders. Since these diseases confer a heavy financial burden on the health care system, AMS detection in point-of-care is fundamental. Here, a one-step assay for direct determination of the AMS activity was developed using a portable personal glucose meter (PGM). In this assay, maltopentaose was used as a substrate for sensitive detection of AMS with the assistance of α-glucosidase. In the presence of AMS, maltopentaose can be readily hydrolyzed to form maltotriose and maltose quickly. With the enzymatic hydrolysis of α-glucosidase, maltotriose and maltose can be turned into glucose rapidly, which can be quantitatively measured using a portable PGM. This assay did not require any cumbersome and time consuming operations, such as surface modification, synthesis of invertase conjugate, washing and centrifugation. Detection of AMS can be achieved using only a one-step mixture, and the limit of detection was 20 U L−1 which was lower than the clinical cutoff for AMS. More importantly, this sensitive and selective assay was also used for the detection of AMS in human serum/urine samples. The results showed that the recovery of AMS from human serum/urine samples was 91–107%. The rapid and easy-to-operate assay may have potential application in the fields of point-of-care (POC) clinical diagnosis, particularly in rural and remote areas where lab equipment may be limited.
Co-reporter:Qing Wang, Xiaohan Yang, Xiaohai Yang, Kemin Wang, Hua Zhang and Pei Liu
Analyst 2015 vol. 140(Issue 22) pp:7657-7662
Publication Date(Web):21 Sep 2015
DOI:10.1039/C5AN01592H
A novel unmodified gold nanoparticle (AuNP)-based colorimetric assay was demonstrated using split aptamers and the hybridization chain reaction (HCR) amplification strategy. Here, the aptamer was divided into a structure-switching hairpin probe (DNA probe H1 (or H1′)) and a single-stranded probe (DNA probe H2 (or H2′)). In the presence of the target, DNA probe H1 (or H1′) could specifically capture the target with the assistance of DNA probe H2 (or H2′) to form a stable complex. Subsequently, the hairpin structure of DNA probe H1 (or H1′) was changed, and then a chain reaction of hybridization events between two other hairpin probes (H3 and H4) propagated, resulting in the formation of nicked double-helices. Since it was difficult for such nicked double-helices to inhibit salt-induced AuNP aggregation, a red-to-blue color change was observed. With the elegant amplification effect of HCR, this assay showed a low detection limit (15 nM for Hg2+ and 1 μM for adenosine), which was lower than or at least comparable to previous AuNP-based methods. The novel strategy not only eliminated the requirements of enzymatic reactions, separation processes, chemical modifications, and sophisticated instruments, but also could be used for other targets only by simply changing the DNA probe sequences.
Co-reporter:Chunlei Wu, Jianbo Liu, Pengfei Zhang, Jing Li, Haining Ji, Xiaohai Yang and Kemin Wang
Analyst 2015 vol. 140(Issue 17) pp:6100-6107
Publication Date(Web):06 Jul 2015
DOI:10.1039/C5AN01145K
A highly specific recognition-before-labeling strategy has been developed for sensitive detection of non-small cell lung cancer A549 cells, by using fluorescent QDs as signal units and DNA aptamers as recognition elements. A QD–aptamer system used for cell imaging and bioanalysis mostly relies on the recognition-after-labeling strategy in which aptamers were firstly labeled with QDs and then the QD–aptamer conjugates as a whole were utilized for specific recognition. Here in our strategy, aptamers were used firstly to recognize target cells, and then fluorescent QDs were sequentially added to bind the aptamers and light the target cells. The proposed recognition-before-labeling strategy didn't require the complex process of QD functionalization, and avoided the possible impact on the aptamer configuration from steric hindrance. Meanwhile, QDs, with strong fluorescence and good photostability, also give this method a high signal-to-background ratio (S/B). The recognition-before-labeling strategy is simple and sensitive, suggesting a new method for in vitro diagnostic assays of cancer cells.
Co-reporter:Xiaochen Guo, Xiaohai Yang, Pei Liu, Kemin Wang, Qing Wang, Qiuping Guo, Jin Huang, Wenshan Li, Fengzhou Xu and Chunxia Song
Analyst 2015 vol. 140(Issue 12) pp:4291-4297
Publication Date(Web):23 Apr 2015
DOI:10.1039/C5AN00626K
MicroRNAs (miRNAs) participate in various biological processes during the course of life. The levels of miRNAs can be useful biomarkers for cellular events or cancer diagnosis, thus sensitive and accurate analysis of miRNA expression is crucial for better understanding its functions and the early diagnosis of human disease. Here, we developed a multiple amplification detection method for miRNA based on the host–guest interaction between β-cyclodextrin polymer and pyrene, which takes advantage of the polymerase-aided strand displacement amplification and λ exonuclease-assisted cyclic enzymatic amplification. The proposed method allowed quantitative detection of miRNA-21 in a dynamic range of 1 pM to 5 nM with a detection limit of 0.3 pM and demonstrated good ability to discriminate the target sequence from the single-base mismatched miRNA sequence. Moreover, the assay was applied successfully in a complex biological matrix. We believe that this proposed sensitive and specific assay has great potential as a quantification method for miRNA detection in biomedical research and clinical diagnosis.
Co-reporter:Qing Wang, Xiaohan Yang, Xiaohai Yang, Fang Liu, Kemin Wang
Sensors and Actuators B: Chemical 2015 212() pp: 440-445
Publication Date(Web):
DOI:10.1016/j.snb.2015.02.040
Co-reporter:Haihua Huang, Xiaohai Yang, Kemin Wang, Qing Wang, Qiuping Guo, Jin Huang, Jianbo Liu, Xiaochen Guo, Wenshan Li, Leiliang He
Talanta 2015 Volume 144() pp:529-534
Publication Date(Web):1 November 2015
DOI:10.1016/j.talanta.2015.06.087
•Our assay is based on the host–guest interaction between β-CDP and pyrene.•The hairpin probe is easy to design, synthesize, purify and convenient for usage.•The detection system relies on hybridization and strand displacement reactions.•Our assay displays fine reproducibility and practical utility in complex samples.The detection of nucleic acids is fundamental for studying their functions and for the development of biological studies and medical diagnostics. Herein, we report a new strategy for nucleic acid amplified detection by combining target-catalyzed dynamic assembly with host–guest interaction between β-cyclodextrin polymer (β-CDP) and pyrene. In this strategy, a metastable pyrene-labeled hairpin DNA probe (probe H1) and a metastable unlabeled hairpin DNA probe (probe H2) were elaborately designed as the assembly components, which were kinetically handicapped from cross-opening in the absence of target DNA. In this state, pyrene labled at the 5ʹ-termini of single-stranded stem of probe H1 would be easily trapped into the hydrophobic cavity of β-CDP because of weak steric hindrance, leading to significant fluorescence enhancement. Once the dynamic assembly was catalyzed by target DNA, a hybridized DNA duplex H1–H2 would be created continuously. In this state, it is difficult for pyrene to enter the cavity of β-CDP due to steric hindrance and weak-binding interaction, leading to a weak fluorescent signal. Thus, target DNA could be detected by this simple mix-and-detect amplification method without the need of expensive and perishable protein enzymes. As low as 10 pM of the target DNA was detected by this assay, which was comparable to that of some reported enzyme-dependent amplification methods. Meanwhile, the proposed method was further successfully applied to detect DNA in cell lysate samples, showing great potential for target detection from complex fluids. In addition, as a novel transformation of dynamic DNA assembly technology into enzyme-free signal-amplification analytical application, the proposed strategy has shown great potential for applications in a wide range of fields, such as aptamer-based non-nucleic acid target sensing, biomedicine and bioimaging.
Co-reporter:Qing Wang, Wei Liu, Yuqian Xing, Xiaohai Yang, Kemin Wang, Rui Jiang, Pei Wang, and Qing Zhao
Analytical Chemistry 2014 Volume 86(Issue 13) pp:6572
Publication Date(Web):June 10, 2014
DOI:10.1021/ac501088q
An aptamer screening method using a positive and negative selection units integrated microfluidic chip was introduced. Here, myoglobin (Myo), one of the early markers to increase after acute myocardial infarction, was used as the model. After 7-round selection, the aptamers, which exhibited dissociation constants (Kd) in the nanomolar range (from 4.93 to 6.38 nM), were successfully obtained using a positive and negative selection units integrated microfluidic chip. The aptamer with the highest affinity (Kd = 4.93 nM) was then used for the fabrication of a label-free supersandwich electrochemical biosensor for Myo detection based on target-induced aptamer displacement. The detection limit of this aptamer-based electrochemical biosensor was 10 pM, which was significantly lower than that of those previous antibody-based biosensors for Myo detection. This work may not only develop a strategy for screening aptamer but also offer promising alternatives to the traditional analytical and immunological methods for Myo detection.
Co-reporter:Chunxia Song, Xiaohai Yang, Kemin Wang, Qing Wang, Jin Huang, Jianbo Liu, Wei Liu, Pei Liu
Analytica Chimica Acta 2014 Volume 827() pp:74-79
Publication Date(Web):27 May 2014
DOI:10.1016/j.aca.2014.04.006
•This method avoided enzyme introduction, operated at mild conditions.•Free of any conjugation or labeling process.•High specificity of target DNA detection against single-base mismatch sequences.A label-free and non-enzymatic amplification fluorescent method for detection of DNA has been developed by using hybridization chain reaction (HCR) and dsDNA-templated copper nanoparticles (CuNPs). First, the biotinylated capture DNA probes were immobilized on the streptavidin-modified beads through the interaction of biotin and streptavidin. Then, target DNA hybridized with the capture DNA probes, which formed a hybridized DNA with sticky end. The sticky end triggered the HCR process and formation of dsDNA polymers while two hairpin probes coexisted. Subsequently, the dsDNA polymers were employed as template for synthesis of CuNPs with excellent fluorescent properties, which provided a label-free, non-enzymatic signal response. Meanwhile, the fluorescence sensing depended on the target DNA triggered HCR, which render this method a high selectivity against single-base mismatch sequences. The concept and methodology developed in this work show great promise in the quantitative detection of DNA in biological and medical applications.A label-free and non-enzymatic amplification fluorescent method for detection of DNA has been developed by using hybridization chain reaction (HCR) and dsDNA-templated copper nanoparticles (CuNPs).
Co-reporter:Pei Liu, Xiao-Hai Yang, Qing Wang, Jing Huang, Jian-Bo Liu, Ying Zhu, Lei-Liang He, Ke-Min Wang
Chinese Chemical Letters 2014 Volume 25(Issue 7) pp:1047-1051
Publication Date(Web):July 2014
DOI:10.1016/j.cclet.2014.05.002
This work develops a fluorescence approach for sensitive detection of DNA methyltransferase activity based on endonuclease and rolling circle amplification (RCA) technique. In the presence of DNA adenine methylation (Dam) MTase, the methylation-responsive sequence of hairpin probe is methylated and cleaved by the methylation-sensitive restriction endonuclease Dpn I. The products cleaved by restriction endonuclease Dpn I then function as a signal primer to initiate RCA reaction by hybridizing with the circular DNA template. Each RCA product containing thousands of repeated sequences might hybridize with a large number of molecular beacons (detection probes), resulting in an enhanced fluorescence signal. In the absence of Dam MTase, neither methylation/cleavage nor RCA reaction can be initiated and no fluorescence signal is observed. The proposed method exhibits a dynamic range from 0.5 U/mL to 30 U/mL and a detection limit of 0.18 U/mL. This method can be used for the screening of antimicrobial drugs and has a great potential to be further applied in early clinical diagnosis.We have developed a simple, sensitive fluorescence method for the detection of MTase activity using hairpin probe-based primer generation rolling circle amplification and molecular beacons.
Co-reporter:Qing Wang;Chenchen Zhou;Lin Liu ;Kemin Wang
Journal of Molecular Recognition 2014 Volume 27( Issue 11) pp:676-682
Publication Date(Web):
DOI:10.1002/jmr.2391
Because cell-specific aptamers have high potential for biomedical applications, investigation of the interaction between cell and its aptamers may be of key importance for an improved understanding of biochemical processes. Herein, the interaction between human lung adenocarcinoma A549 cell and its four aptamers was explored using single-molecule force spectroscopy (SMFS). The values of the unbinding force varied from 117.1 to 171.0 pN at the loading rate of 1.8 × 105 pN/s. Based on the dependence of singe molecule force on the atomic force microscopy loading rate, the corresponding kinetic parameters were obtained. The results revealed two activation barriers and two transient states in the unbinding process of aptamer/cell interaction. More importantly, the binding sites on A549 cells with its four aptamers were defined to be different using SMFS and flow cytometry. This work demonstrated that SMFS can be used as a powerful tool for exploring the aptamer/cell binding behavior at the single-molecule level, and may provide valuable information for the design and application of aptamer probes. Copyright © 2014 John Wiley & Sons, Ltd.
Co-reporter:Jianbo Liu, Yan Liu, Xiaohai Yang, Kemin Wang, Qing Wang, Hui Shi, and Li Li
Analytical Chemistry 2013 Volume 85(Issue 22) pp:11121
Publication Date(Web):October 10, 2013
DOI:10.1021/ac403023p
A novel exciton energy transfer-based ultrasensitive fluorescent sensing strategy for the detection of biological small molecules and protein has been established through split aptamer-programmed self-assembly of quantum dots (QDs). The signal is produced from exciton energy transfer of the self-assembled QDs. The recognition is accomplished using an aptamer sensor scaffold designed with two split fragment sequences, which specifically bind to the model analytes. The extent of particle assembly, induced by the analyte-triggered self-assembly of QDs, leads to an exciton energy transfer effect between interparticles, giving a readily detectable fluorescent quenching and red shift of the emission peak, which enables us to quantitate the target in dual signal modes. The application of the technique is well demonstrated using two representative split aptamer-based model systems for the detection of adenosine and thrombin. The sensitivity of this exciton energy transfer-based fluorescent sensing is much better than that of plasmonic coupling-based colorimetric methods. Limit of detections (LODs) down to 12 nM and 15 pM can be achieved for adenosine and thrombin, respectively. The sensing strategy is proposed as a general platform for robust and specific aptamer–target analysis which could be further developed to monitor a wide range of target analytes. The concept and methodology developed in this work shows a good promise in the study of molecular binding events in the biological and medical applications.
Co-reporter:Wenshan Li, Lei Luo, Jin Huang, Qing Wang, Jianbo Liu, Xiao Xiao, Hongmei Fang, Xiaohai Yang and Kemin Wang
Chemical Communications 2017 - vol. 53(Issue 40) pp:NaN5568-5568
Publication Date(Web):2017/04/18
DOI:10.1039/C7CC01128H
We report self-assembled nanocentipedes as multivalent vehicles for the delivery of immunostimulatory CpG ODNs. The multivalent vehicles could be internalized by RAW264.7 cells and stimulate the secretion of large amounts of cytokines, successively inducing effective apoptosis of cancer cells.
Co-reporter:Min Lin, Wenshan Li, Yaning Wang, Xiaohai Yang, Kemin Wang, Qing Wang, Pei Wang, Yujie Chang and Yuyu Tan
Chemical Communications 2015 - vol. 51(Issue 39) pp:NaN8306-8306
Publication Date(Web):2015/04/09
DOI:10.1039/C5CC00929D
Discrimination of hemoglobins with subtle differences was achieved using an aptamer based sensing array. Linear discriminant analysis (LDA) showed that the sensing array can discriminate human hemoglobins from hemoglobins of different species.
Co-reporter:Chunxia Song, Xiaohai Yang, Kemin Wang, Qing Wang, Jianbo Liu, Jin Huang, Leiliang He, Pei Liu, Zhihe Qing and Wei Liu
Chemical Communications 2015 - vol. 51(Issue 10) pp:NaN1818-1818
Publication Date(Web):2014/12/10
DOI:10.1039/C4CC08991J
A strategy for T4 polynucleotide kinase activity detection was proposed based on a β-cyclodextrin polymer (polyβ-CD) and an exonuclease reaction. The fluorescence of pyrene enhanced by more than 10 times in the presence of polyβ-CD, and a simple detection of T4 PNK was achieved with a detection limit of 0.02 units per mL.
Co-reporter:Nandi Chen, Xiaohai Yang, Qing Wang, Lixin Jian, Hui Shi, Shiya Qin, Kemin Wang, Jin Huang and Wenjing Liu
Chemical Communications 2016 - vol. 52(Issue 41) pp:NaN6792-6792
Publication Date(Web):2016/04/20
DOI:10.1039/C6CC02374F
Existing drug delivery systems were not suitable for killing cells in the circulatory system specifically. Herein, we developed a novel localized drug delivery strategy, in which the release of anticancer agents was specifically triggered by circulating tumor cells. Meanwhile, damage to non-target cells was avoided.
Co-reporter:Bin Wu, Rui Jiang, Qing Wang, Jin Huang, Xiaohai Yang, Kemin Wang, Wenshan Li, Nandi Chen and Qing Li
Chemical Communications 2016 - vol. 52(Issue 17) pp:NaN3571-3571
Publication Date(Web):2016/01/27
DOI:10.1039/C5CC10486F
High affinity DNA aptamers against C-reactive protein (CRP) were obtained using a microfluidic chip. The aptamers were then used for the construction of an Au nanoparticle enhanced surface plasmon resonance biosensor, which was introduced for the detection of CRP at concentrations ranging from 10 pM to 100 nM in diluted human serum.
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