Co-reporter:Tian Tian, Yuan An, Yiping Wu, Yanling Song, Zhi Zhu, and Chaoyong Yang
ACS Applied Materials & Interfaces September 13, 2017 Volume 9(Issue 36) pp:30480-30480
Publication Date(Web):August 17, 2017
DOI:10.1021/acsami.7b09717
An integrated distance-based origami paper analytical device (ID-oPAD) is developed for simple, user friendly and visual detection of targets of interest. The platform enables complete integration of target recognition, signal amplification, and visual signal output based on aptamer/invertase-functionalized sepharose beads, cascaded enzymatic reactions, and a 3D microfluidic paper-based analytical device with distance-based readout, respectively. The invertase–DNA conjugate is released upon target addition, after which it permeates through the cellulose and flows down into the bottom detection zone, whereas sepharose beads with larger size are excluded and stay in the upper zone. Finally, the released conjugate initiates cascaded enzymatic reactions and translates the target signal into a brown bar chart reading. By simply closing the device, the ID-oPAD enables a sample-in-answer-out assay within 30 min with visual and quantitative readout. Importantly, bound/free probe separation is achieved by taking advantage of the size difference between sepharose beads and cellulose pores, and the downstream enzymatic amplification is realized based on the compatibility of multiple enzymes with corresponding substrates. Overall, with the advantages of low-cost, disposability, simple operation, and visual quantitative readout, the ID-oPAD offers an ideal platform for point-of-care testing, especially in resource-limited areas.Keywords: aptamer; distance-based detection; microfluidic paper-based devices; point-of-care (POC);
Co-reporter:Dan Liu, Shasha Jia, Huimin Zhang, Yanli Ma, Zhichao Guan, Jiuxing Li, Zhi Zhu, Tianhai Ji, and Chaoyong James Yang
ACS Applied Materials & Interfaces July 12, 2017 Volume 9(Issue 27) pp:22252-22252
Publication Date(Web):June 26, 2017
DOI:10.1021/acsami.7b05531
Point-of-care testing (POCT) with the advantages of speed, simplicity, and low cost, as well as no need for instrumentation, is critical for the measurement of analytes in a variety of environments lacking access to laboratory infrastructure. In the present study, a hydrogel pressure-based assay for quantitative POCT was developed by integrating a target-responsive hydrogel with pressuremeter readout. The target-responsive hydrogels were constructed with DNA grafted linear polyacrylamide and the cross-linking DNA for selective target recognition. The hydrogel response to the target substance allows release of the preloaded Pt nanoparticles, which have good stability and excellent catalytic ability for decomposing H2O2 to O2. Then, the generated O2 in a sealed environment leads to significant pressure increase, which can be easily read out by a handheld pressuremeter. Using this target-responsive hydrogel pressure-based assay, portable and highly sensitive detection of cocaine, ochratoxin A, and lead ion were achieved with excellent accuracy and selectivity. With the advantages of portability, high sensitivity, and simple sample processing, the hydrogel pressure-based assay shows great potential for quantitative POCT of a broad range of targets in resource-limited settings.Keywords: gas generation; pressuremeter readout; Pt nanoparticles; quantitative point-of-care testing; target-responsive hydrogel;
Co-reporter:Zhi ZhuChaoyong James Yang
Accounts of Chemical Research 2017 Volume 50(Issue 1) pp:
Publication Date(Web):December 28, 2016
DOI:10.1021/acs.accounts.6b00370
ConspectusHeterogeneity among individual molecules and cells has posed significant challenges to traditional bulk assays, due to the assumption of average behavior, which would lose important biological information in heterogeneity and result in a misleading interpretation. Single molecule/cell analysis has become an important and emerging field in biological and biomedical research for insights into heterogeneity between large populations at high resolution. Compared with the ensemble bulk method, single molecule/cell analysis explores the information on time trajectories, conformational states, and interactions of individual molecules/cells, all key factors in the study of chemical and biological reaction pathways. Various powerful techniques have been developed for single molecule/cell analysis, including flow cytometry, atomic force microscopy, optical and magnetic tweezers, single-molecule fluorescence spectroscopy, and so forth. However, some of them have the low-throughput issue that has to analyze single molecules/cells one by one. Flow cytometry is a widely used high-throughput technique for single cell analysis but lacks the ability for intercellular interaction study and local environment control. Droplet microfluidics becomes attractive for single molecule/cell manipulation because single molecules/cells can be individually encased in monodisperse microdroplets, allowing high-throughput analysis and manipulation with precise control of the local environment. Moreover, hydrogels, cross-linked polymer networks that swell in the presence of water, have been introduced into droplet microfluidic systems as hydrogel droplet microfluidics. By replacing an aqueous phase with a monomer or polymer solution, hydrogel droplets can be generated on microfluidic chips for encapsulation of single molecules/cells according to the Poisson distribution. The sol–gel transition property endows the hydrogel droplets with new functionalities and diversified applications in single molecule/cell analysis. The hydrogel can act as a 3D cell culture matrix to mimic the extracellular environment for long-term single cell culture, which allows further heterogeneity study in proliferation, drug screening, and metastasis at the single-cell level. The sol–gel transition allows reactions in solution to be performed rapidly and efficiently with product storage in the gel for flexible downstream manipulation and analysis. More importantly, controllable sol–gel regulation provides a new way to maintain phenotype-genotype linkages in the hydrogel matrix for high throughput molecular evolution. In this Account, we will review the hydrogel droplet generation on microfluidics, single molecule/cell encapsulation in hydrogel droplets, as well as the progress made by our group and others in the application of hydrogel droplet microfluidics for single molecule/cell analysis, including single cell culture, single molecule/cell detection, single cell sequencing, and molecular evolution.
Co-reporter:Yishun Huang;Xuemeng Wu;Tian Tian;Zhi Zhu;Hui Lin
Science China Chemistry 2017 Volume 60( Issue 2) pp:293-298
Publication Date(Web):2017 February
DOI:10.1007/s11426-016-0242-2
Lanthanide elements (Ln) play an important role in industry and agriculture. As a result of the increasing consumption of lanthanides, environmental emission of Ln has become detrimental to the health of flora and fauna. Current methods for trace lanthanides detection mainly rely on sophisticated instruments. In this article, a Ln3+ dependent DNAzyme was incorporated into a hydrogel to generate Ln3+ sensitive DNAzyme hydrogel for portable colorimetric detection. The enzyme strand and its substrate strand act as crosslinker and functional unit of the hydrogel with polyacrylamide chains as the scaffold and gold nanoparticles (AuNPs) as the indicator of hydrogel stability. Any ions in the Ln3+ series can trigger the cleavage of substrate strand by activating the enzyme strand, thereby decreasing the crosslink ratio and leading to collapse of the hydrogel. The release of the encapsulated AuNPs turns the supernatant wine red. Using this colorimetric method, Ln3+ can be detected with high sensitivity, with a limit of detection (LOD) of 20 nM for Ce3+. The hydrogel responds specifically to any Ln3+ ion and works well with the spiked lake sample without the need of instruments and skilled operators. Our results suggest that the lanthanide responsive hydrogel can be used for portable and sensitive detection of Ln3+ contamination in the field.
Co-reporter:Yanling Song;Yuan An;Weizhi Liu;Wanfu Hou;Xingrui Li;Bingqian Lin;Zhi Zhu;Shengxiang Ge;Huang-hao Yang;Chaoyong Yang
Chemical Communications 2017 vol. 53(Issue 86) pp:11774-11777
Publication Date(Web):2017/10/26
DOI:10.1039/C7CC07231G
This paper describes a centrifugal micropipette-tip method for ELISA sample processing combined with a pressure meter for portable quantitative detection of acute myocardial infarction (AMI) biomarker myoglobin (Myo). The method enables sensitive and reliable quantification of Myo in 35 minutes. With the advantages of simplicity, speed, user friendliness, low cost, and small sample consumption, centrifugal micropipette-tip ELISA shows great potential for quantitative POC diagnostics for AMI.
Co-reporter:Yanling Song;Tian Tian;Yuanzhi Shi;Wenli Liu;Yuan Zou;Tahereh Khajvand;Sili Wang;Zhi Zhu;Chaoyong Yang
Chemical Science (2010-Present) 2017 vol. 8(Issue 3) pp:1736-1751
Publication Date(Web):2017/02/28
DOI:10.1039/C6SC04671A
Up to 90% of cancer-related deaths are caused by metastatic cancer. Circulating tumor cells (CTCs), a type of cancer cell that spreads through the blood after detaching from a solid tumor, are essential for the establishment of distant metastasis for a given cancer. As a new type of liquid biopsy, analysis of CTCs offers the possibility to avoid invasive tissue biopsy procedures with practical implications for diagnostics. The fundamental challenges of analyzing and profiling CTCs are the extremely low abundances of CTCs in the blood and the intrinsic heterogeneity of CTCs. Various technologies have been proposed for the enrichment and single-cell analysis of CTCs. This review aims to provide in-depth insights into CTC analysis, including various techniques for isolation of CTCs with capture methods based on physical and biochemical principles, and single-cell analysis of CTCs at the genomic, proteomic and phenotypic level, as well as current developmental trends and promising research directions.
Co-reporter:Dan Liu, Xingrui Li, Junkai Zhou, Shibo Liu, Tian Tian, Yanling Song, Zhi Zhu, Leiji Zhou, Tianhai Ji, Chaoyong Yang
Biosensors and Bioelectronics 2017 Volume 96(Volume 96) pp:
Publication Date(Web):15 October 2017
DOI:10.1016/j.bios.2017.04.044
•We constructed a fully integrated ELISA-Chip with sample-in-answer-out capability for POCT.•.The molecular recognition event of ELISA was converted into amplified gas-generation reaction with distance readout.•Our ELISA-Chip allows ultrasensitive quantitation of disease biomarkers within 2 h without additional equipment.•The ELISA-Chip is inexpensive, disposable and highly suitable for POCT in resource-limited settings.Enzyme-linked immunosorbent assay (ELISA) is a popular laboratory technique for detection of disease-specific protein biomarkers with high specificity and sensitivity. However, ELISA requires labor-intensive and time-consuming procedures with skilled operators and spectroscopic instrumentation. Simplification of the procedures and miniaturization of the devices are crucial for ELISA-based point-of-care (POC) testing in resource-limited settings. Here, we present a fully integrated, instrument-free, low-cost and portable POC platform which integrates the process of ELISA and the distance readout into a single microfluidic chip. Based on manipulation using a permanent magnet, the process is initiated by moving magnetic beads with capture antibody through different aqueous phases containing ELISA reagents to form bead/antibody/antigen/antibody sandwich structure, and finally converts the molecular recognition signal into a highly sensitive distance readout for visual quantitative bioanalysis. Without additional equipment and complicated operations, our integrated ELISA-Chip with distance readout allows ultrasensitive quantitation of disease biomarkers within 2 h. The ELISA-Chip method also showed high specificity, good precision and great accuracy. Furthermore, the ELISA-Chip system is highly applicable as a sandwich-based platform for the detection of a variety of protein biomarkers. With the advantages of visual analysis, easy operation, high sensitivity, and low cost, the integrated sample-in-answer-out ELISA-Chip with distance readout shows great potential for quantitative POCT in resource-limited settings.
Co-reporter:Metages Gashaw Ahmed;Mahlet Fasil Abate;Dr. Yanling Song; Zhi Zhu; Feng Yan;Yao Xu; Xiaomin Wang; Qingbiao Li; Chaoyong Yang
Angewandte Chemie International Edition 2017 Volume 56(Issue 36) pp:10681-10685
Publication Date(Web):2017/08/28
DOI:10.1002/anie.201702675
AbstractEven though the diagnostic and prognostic value of circulating tumor cells (CTCs) has been demonstrated, their clinical utility and widespread adoption have been limited. Herein, we describe a new device, size-dictated immunocapture chip (SDI-Chip), for efficient, sensitive, and spatially resolved capture and detection of CTCs. SDI-Chip enables selective, frequent, and extended interaction of CTCs with hydrodynamically optimized immunocoated micropillar surfaces. CTCs with different antigen expression levels can be efficiently captured and spatially resolved around the micropillars. Capture efficiency greater than 92 % with a purity of 82 % was achieved with blood samples. CTCs were detected in non-metastasis colorectal (CRC) patients, while none was detected from healthy volunteers. We believe that SDI-Chip will facilitate the transition of tumor diagnosis from anatomical pathology to molecular pathology in localized CRC patients.
Co-reporter:Bingqian Lin, Dan Liu, Jinmao Yan, Zhi Qiao, Yunxin Zhong, Jiawei Yan, Zhi Zhu, Tianhai Ji, and Chaoyong James Yang
ACS Applied Materials & Interfaces 2016 Volume 8(Issue 11) pp:6890
Publication Date(Web):February 26, 2016
DOI:10.1021/acsami.6b00777
There is considerable demand for sensitive, selective, and portable detection of disease-associated proteins, particularly in clinical practice and diagnostic applications. Portable devices are highly desired for detection of disease biomarkers in daily life due to the advantages of being simple, rapid, user-friendly, and low-cost. Herein we report an enzyme-encapsulated liposome-linked immunosorbent assay for sensitive detection of proteins using personal glucose meters (PGM) for portable quantitative readout. Liposomes encapsulating a large amount of amyloglucosidase or invertase are surface-coated with recognition elements such as aptamers or antibodies for target recognition. By translating molecular recognition signal into a large amount of glucose with the encapsulated enzyme, disease biomarkers such as thrombin or C-reactive protein (CRP) can be quantitatively detected by a PGM with a high detection limit of 1.8 or 0.30 nM, respectively. With the advantages of portability, ease of use, and low-cost, the method reported here has potential for portable and quantitative detection of various targets for different POC testing scenarios, such as rapid diagnosis in clinic offices, health monitoring at the bedside, and chemical/biochemical safety control in the field.Keywords: disease biomarkers; enzyme-encapsulated liposome; personal glucose meter; portable detection; signal amplification
Co-reporter:Tian Tian, Jiuxing Li, Yanling Song, Leiji Zhou, Zhi Zhu and Chaoyong James Yang
Lab on a Chip 2016 vol. 16(Issue 7) pp:1139-1151
Publication Date(Web):18 Feb 2016
DOI:10.1039/C5LC01562F
Equipment-free devices with quantitative readout are of great significance to point-of-care testing (POCT), which provides real-time readout to users and is especially important in low-resource settings. Among various equipment-free approaches, distance-based visual quantitative detection methods rely on reading the visual signal length for corresponding target concentrations, thus eliminating the need for sophisticated instruments. The distance-based methods are low-cost, user-friendly and can be integrated into portable analytical devices. Moreover, such methods enable quantitative detection of various targets by the naked eye. In this review, we first introduce the concept and history of distance-based visual quantitative detection methods. Then, we summarize the main methods for translation of molecular signals to distance-based readout and discuss different microfluidic platforms (glass, PDMS, paper and thread) in terms of applications in biomedical diagnostics, food safety monitoring, and environmental analysis. Finally, the potential and future perspectives are discussed.
Co-reporter:Yanling Song, Yuanzhi Shi, Xingrui Li, Yanli Ma, Mingxuan Gao, Dan Liu, Yu Mao, Zhi Zhu, Hui Lin, and Chaoyong Yang
Analytical Chemistry 2016 Volume 88(Issue 16) pp:8294
Publication Date(Web):July 25, 2016
DOI:10.1021/acs.analchem.6b02140
Binding affinity characterization is of great importance for aptamer screening because the dissociation constant (Kd) value is a key parameter for evaluating molecular interaction. However, conventional methods often require sophisticated equipment and time-consuming processing. Here, we present a portable device, Afi-Chip, as an equipment-free, rapid, low-cost, and universal platform for evaluation of the aptamer affinity. The Afi-Chip displays a distance readout based on the reaction of an enzyme catalyzing the decomposition of H2O2 for gas generation to push the movement of ink bar. Taking advantage of translating the recognition signal to distance signal and realizing the regents mixing and quantitative readout on the chip, we successfully monitored the aptamer evolution process and characterized binding affinity of aptamers against multiple types of targets, including small molecule glucose, cancer biomarker protein EpCAM, and tumor cell SW620. We also applied the Afi-Chip for rapid characterization of the affinity between anti-HCG and HCG to demonstrate the generality for the molecular interaction study. All of the Kd values obtained are comparable to those reported in the literature or obtained by sophisticated instruments such as a flow cytometer. The Afi-Chip offers a new approach for equipment-free investigation of molecular interactions, such as aptamer identification, ligand selection monitoring, and drug screening.
Co-reporter:Jiuxing Li, Zhi Zhu, Bingqing Zhu, Yanli Ma, Bingqian Lin, Rudi Liu, Yanling Song, Hui Lin, Song Tu, and Chaoyong Yang
Analytical Chemistry 2016 Volume 88(Issue 15) pp:7828
Publication Date(Web):July 6, 2016
DOI:10.1021/acs.analchem.6b01867
Due to its large enhancement effect, nanostructure-based surface-enhanced Raman scattering (SERS) technology had been widely applied for bioanalysis and cell imaging. However, most SERS nanostructures suffer from poor signal reproducibility, which hinders the application of SERS nanostructures in quantitative detection. We report an etching-assisted approach to synthesize SERS-active plasmonic nanoparticles with 1 nm interior nanogap for multiplex quantitative detection and cancer cell imaging. Raman dyes and methoxy poly(ethylene glycol) thiol (mPEG–SH) were attached to gold nanoparticles (AuNPs) to prepare gold cores. Next, Ag atoms were deposited on gold cores in the presence of Pluronic F127 to form a Ag shell. HAuCl4 was used to etch the Ag shell and form an interior nanogap in Au@AgAuNPs, leading to increased Raman intensity of dyes. SERS intensity distribution of Au@AgAuNPs was found to be more uniform than that of aggregated AuNPs. Finally, Au@AgAuNPs were used for multiplex quantitative detection and cancer cell imaging. With the advantages of simple and rapid preparation of Au@AgAuNPs with highly uniform, stable, and reproducible Raman intensity, the method reported here will widen the applications of SERS-active nanoparticles in diagnostics and imaging.
Co-reporter:Xiaofeng Wei, Tian Tian, Shasha Jia, Zhi Zhu, Yanli Ma, Jianjun Sun, Zhenyu Lin, and Chaoyong James Yang
Analytical Chemistry 2016 Volume 88(Issue 4) pp:2345
Publication Date(Web):January 14, 2016
DOI:10.1021/acs.analchem.5b04294
A disposable, equipment-free, versatile point-of-care testing platform, microfluidic distance readout sweet hydrogel integrated paper-based analytical device (μDiSH-PAD), was developed for portable quantitative detection of different types of targets. The platform relies on a target-responsive aptamer cross-linked hydrogel for target recognition, cascade enzymatic reactions for signal amplification, and microfluidic paper-based analytic devices (μPADs) for visual distance-based quantitative readout. A “sweet” hydrogel with trapped glucoamylase (GA) was synthesized using an aptamer as a cross-linker. When target is present in the sample, the “sweet” hydrogel collapses and releases enzyme GA into the sample, generating glucose by amylolysis. A hydrophilic channel on the μPADs is modified with glucose oxidase (GOx) and colorless 3,3′-diaminobenzidine (DAB) as the substrate. When glucose travels along the channel by capillary action, it is converted to H2O2 by GOx. In addition, DAB is converted into brown insoluble poly-3,3′-diaminobenzidine [poly(DAB)] by horseradish peroxidase, producing a visible brown bar, whose length is positively correlated to the concentration of targets. The distance-based visual quantitative platform can detect cocaine in urine with high selectivity, sensitivity, and accuracy. Because the target-induced cascade reaction is triggered by aptamer/target recognition, this method is widely suitable for different kinds of targets. With the advantages of low cost, ease of operation, general applicability, and disposability with quantitative readout, the μDiSH-PAD holds great potential for portable detection of trace targets in environmental monitoring, security inspection, personalized healthcare, and clinical diagnostics.
Co-reporter:Weiting Zhang, Xiaolong Zu, Yanling Song, Zhi Zhu and Chaoyong James Yang
Analyst 2016 vol. 141(Issue 2) pp:579-584
Publication Date(Web):05 Oct 2015
DOI:10.1039/C5AN01763G
Abnormal DNA methylation patterns caused by altered DNA methyltransferase (MTase) activity are closely associated with cancer. Herein, using DNA adenine methylation methyltransferase (Dam MTase) as a model analyte, we designed an allosteric molecular beacon (aMB) for sensitive detection of Dam MTase activity. When the specific site in an aMB is methylated by Dam MTase, the probe can be cut by the restriction nuclease DpnI to release a fluorophore labeled aptamer specific for streptavidin (SA) which will bind to SA beads to generate highly fluorescent beads for easy signal readout by a microscope or flow cytometer. However, aMBs maintain a hairpin structure without the binding ability to SA beads in the absence of Dam MTase, leading to weakly fluorescent SA beads. Unlike the existing signal amplified assays, our method is simpler and more convenient. The high performance of the aptamer and the easy bead separation process make this probe superior to other methods for the detection of MTase in complex biological systems. Overall, the proposed method with a detection limit of 0.57 U mL−1 for Dam MTase shows great potential for further applications in the detection of other MTases, screening of MTase inhibitors, and early diagnosis of cancer.
Co-reporter:Yishun Huang, Luting Fang, Zhi Zhu, Yanli Ma, Leiji Zhou, Xi Chen, Dunming Xu, Chaoyong Yang
Biosensors and Bioelectronics 2016 Volume 85() pp:496-502
Publication Date(Web):15 November 2016
DOI:10.1016/j.bios.2016.05.008
•We constructed a DNAzyme hydrogel system for portable detection of Uranium.•UO22+ dependent DNAzyme crosslinked hydrogel was designed and synthesized.•By encapsulating AuNPs inside hydrogel, UO22+ can be visually detected by naked eyes.•Portable quantitative detection was achieved by encapsulating PtNPs with V-Chip.Due to uranium's increasing exploitation in nuclear energy and its toxicity to human health, it is of great significance to detect uranium contamination. In particular, development of a rapid, sensitive and portable method is important for personal health care for those who frequently come into contact with uranium ore mining or who investigate leaks at nuclear power plants. The most stable form of uranium in water is uranyl ion (UO22+). In this work, a UO22+ responsive smart hydrogel was designed and synthesized for rapid, portable, sensitive detection of UO22+. A UO22+ dependent DNAzyme complex composed of substrate strand and enzyme strand was utilized to crosslink DNA-grafted polyacrylamide chains to form a DNA hydrogel. Colorimetric analysis was achieved by encapsulating gold nanoparticles (AuNPs) in the DNAzyme-crosslinked hydrogel to indicate the concentration of UO22+. Without UO22+, the enzyme strand is not active. The presence of UO22+ in the sample activates the enzyme strand and triggers the cleavage of the substrate strand from the enzyme strand, thereby decreasing the density of crosslinkers and destabilizing the hydrogel, which then releases the encapsulated AuNPs. As low as 100 nM UO22+ was visually detected by the naked eye. The target-responsive hydrogel was also demonstrated to be applicable in natural water spiked with UO22+. Furthermore, to avoid the visual errors caused by naked eye observation, a previously developed volumetric bar-chart chip (V-Chip) was used to quantitatively detect UO22+ concentrations in water by encapsulating Au-Pt nanoparticles in the hydrogel. The UO22+ concentrations were visually quantified from the travelling distance of ink-bar on the V-Chip. The method can be used for portable and quantitative detection of uranium in field applications without skilled operators and sophisticated instruments.
Co-reporter:Tian Tian, Xiaofeng Wei, Shasha Jia, Ruihua Zhang, Jiuxing Li, Zhi Zhu, Huimin Zhang, Yanli Ma, Zhenyu Lin, Chaoyong James Yang
Biosensors and Bioelectronics 2016 Volume 77() pp:537-542
Publication Date(Web):15 March 2016
DOI:10.1016/j.bios.2015.09.049
•We constructed a hydrogel–μPAD system for POCT.•A glucoamylase-trapped aptamer-crosslinked hydrogel was used for molecular recognition.•Cascade enzymatic reactions were integrated for signal amplification•The system allows semiquantitation by naked eyes and quantitation by handheld devicesµPADs were used for portable readout.Paper based microfluidics (µPADs) with advantages of portability, low cost, and ease of use have attracted extensive attention. Here we describe a novel method that integrates glucoamylase-trapped aptamer-crosslinked hydrogel for molecular recognition with cascaded enzymatic reactions for signal amplification and a µPAD for portable readout. Upon target introduction, the hydrogel decomposes to release glucoamylase, which catalyzes the hydrolysis of amylose to produce a large amount of glucose. With a simple folding of the µPAD, the sample solution containing glucose product wicks and diffuses in parallel to each test-zone to carry out homogeneous assays, where glucose is used to produce I2 for brown color visualization through multiple enzymatic and chemical cascade reactions. Through color gradient changes based on different concentrations of the target, a semiquantitative assay is achieved by the naked eye, and quantitation can be obtained by handheld devices. Detection of cocaine in buffer and urine was performed to demonstrate the utility of the hydrogel–µPAD system. More importantly, the hydrogel–µPAD system can be extended to the detection of various targets by incorporating the corresponding aptamer into the hydrogel. The hydrogel–µPAD system reported here provides a new platform for portable, disposable and visual detection of a wide range of targets.
Co-reporter:B.Sc. Huimin Zhang; Leiji Zhou; Zhi Zhu ; Chaoyong Yang
Chemistry - A European Journal 2016 Volume 22( Issue 29) pp:9886-9900
Publication Date(Web):
DOI:10.1002/chem.201503543
Abstract
Nucleic acid aptamers are short synthetic DNA or RNA sequences that can bind to a wide range of targets with high affinity and specificity. In recent years, aptamers have attracted increasing research interest due to their unique features of high binding affinity and specificity, small size, excellent chemical stability, easy chemical synthesis, facile modification, and minimal immunogenicity. These properties make aptamers ideal recognition ligands for bioanalysis, disease diagnosis, and cancer therapy. This review highlights the recent progress in aptamer selection and the latest applications of aptamer-based functional probes in the fields of bioanalysis and biomedicine.
Co-reporter:Yi Xie, Xiaoyan Lin, Yishun Huang, Rujun Pan, Zhi Zhu, Leiji Zhou and Chaoyong James Yang
Chemical Communications 2015 vol. 51(Issue 11) pp:2156-2158
Publication Date(Web):17 Dec 2014
DOI:10.1039/C4CC08912J
Based on the protective properties of polydopamine nanospheres for DNA probes against nuclease digestion, we have developed a DNase I-assisted target recycling signal amplification method for highly sensitive and selective detection of miRNA.
Co-reporter:Rudi Liu, Yishun Huang, Yanli Ma, Shasha Jia, Mingxuan Gao, Jiuxing Li, Huimin Zhang, Dunming Xu, Min Wu, Yan Chen, Zhi Zhu, and Chaoyong Yang
ACS Applied Materials & Interfaces 2015 Volume 7(Issue 12) pp:6982
Publication Date(Web):March 16, 2015
DOI:10.1021/acsami.5b01120
A target-responsive aptamer-cross-linked hydrogel was designed and synthesized for portable and visual quantitative detection of the toxin Ochratoxin A (OTA), which occurs in food and beverages. The hydrogel network forms by hybridization between one designed DNA strand containing the OTA aptamer and two complementary DNA strands grafting on linear polyacrylamide chains. Upon the introduction of OTA, the aptamer binds with OTA, leading to the dissociation of the hydrogel, followed by release of the preloaded gold nanoparticles (AuNPs), which can be observed by the naked eye. To enable sensitive visual and quantitative detection, we encapsulated Au@Pt core–shell nanoparticles (Au@PtNPs) in the hydrogel to generate quantitative readout in a volumetric bar-chart chip (V-Chip). In the V-Chip, Au@PtNPs catalyzes the oxidation of H2O2 to generate O2, which induces movement of an ink bar to a concentration-dependent distance for visual quantitative readout. Furthermore, to improve the detection limit in complex real samples, we introduced an immunoaffinity column (IAC) of OTA to enrich OTA from beer. After the enrichment, as low as 1.27 nM (0.51 ppb) OTA can be detected by the V-Chip, which satisfies the test requirement (2.0 ppb) by the European Commission. The integration of a target-responsive hydrogel with portable enrichment by IAC, as well as signal amplification and quantitative readout by a simple microfluidic device, offers a new method for portable detection of food safety hazard toxin OTA.Keywords: DNA hydrogel; microfluidics; Ochratoxin A; visual detection
Co-reporter:Xiaofeng Wei, Tian Tian, Shasha Jia, Zhi Zhu, Yanli Ma, Jianjun Sun, Zhenyu Lin, and Chaoyong James Yang
Analytical Chemistry 2015 Volume 87(Issue 8) pp:4275
Publication Date(Web):March 25, 2015
DOI:10.1021/acs.analchem.5b00532
A versatile point-of-care assay platform was developed for simultaneous detection of multiple targets based on a microfluidic paper-based analytic device (μPAD) using a target-responsive hydrogel to mediate fluidic flow and signal readout. An aptamer-cross-linked hydrogel was used as a target-responsive flow regulator in the μPAD. In the absence of a target, the hydrogel is formed in the flow channel, stopping the flow in the μPAD and preventing the colored indicator from traveling to the final observation spot, thus yielding a “signal off” readout. In contrast, in the presence of a target, no hydrogel is formed because of the preferential interaction of target and aptamer. This allows free fluidic flow in the μPAD, carrying the indicator to the observation spot and producing a “signal on” readout. The device is inexpensive to fabricate, easy to use, and disposable after detection. Testing results can be obtained within 6 min by the naked eye via a simple loading operation without the need for any auxiliary equipment. Multiple targets, including cocaine, adenosine, and Pb2+, can be detected simultaneously, even in complex biological matrices such as urine. The reported method offers simple, low cost, rapid, user-friendly, point-of-care testing, which will be useful in many applications.
Co-reporter:Xilan Li, Yuan An, Jiang Jin, Zhi Zhu, Linlin Hao, Lu Liu, Yongquan Shi, Daiming Fan, Tianhai Ji, and Chaoyong James Yang
Analytical Chemistry 2015 Volume 87(Issue 9) pp:4941
Publication Date(Web):April 13, 2015
DOI:10.1021/acs.analchem.5b00637
Metastasis, the capability of tumor cells to spread and grow at distant sites, is the primary factor in cancer mortality. Because metastasis in sentinel lymph nodes suggests the original spread of tumors from a primary site, the detection of lymph node involvement with cancer serves as an important prognostic and treatment parameter. Here we have developed a panel of DNA aptamers specifically binding to colon cancer cell SW620 derived from metastatic site lymph node, with high affinity after 14 rounds of selection by the cell-SELEX (systematic evolution of ligands by exponential enrichment) method. The binding affinities of selected aptamers were evaluated by flow cytometry. Aptamer XL-33 with the best binding affinity (0.7 nM) and its truncated sequence XL-33-1 with 45 nt showed excellent selectivity for recognizing target cell SW620. The binding entity of the selected aptamer has been preliminarily determined as a membrane protein on the cell surface. Tissue imaging results showed that XL-33-1 was highly specific to the metastatic tumor tissue or lymph node tissue with corresponding cancer metastasis and displayed an 81.7% detection rate against colon cancer tissue with metastasis in regional lymph nodes. These results suggest that XL-33-1 has great potential to become a molecular imaging agent for early detection of lymph node tissue with colon cancer metastasis. More importantly, this study clearly demonstrates that DNA ligands selectively recognizing metastatic cancer cells can be readily generated by metastatic-cell-based SELEX for potential applications in metastatic cancer diagnosis and treatment.
Co-reporter: Zhi Zhu;Zhichao Guan;Dan Liu;Shasha Jia;Jiuxing Li;Zhichao Lei;Shuichao Lin;Tianhai Ji; Zhongqun Tian ; Chaoyong James Yang
Angewandte Chemie International Edition 2015 Volume 54( Issue 36) pp:
Publication Date(Web):
DOI:10.1002/anie.201583661
Co-reporter: Zhi Zhu;Zhichao Guan;Dan Liu;Shasha Jia;Jiuxing Li;Zhichao Lei;Shuichao Lin;Tianhai Ji; Zhongqun Tian ; Chaoyong James Yang
Angewandte Chemie 2015 Volume 127( Issue 36) pp:
Publication Date(Web):
DOI:10.1002/ange.201583661
Co-reporter: Zhi Zhu;Zhichao Guan;Dan Liu;Shasha Jia;Jiuxing Li;Zhichao Lei;Shuichao Lin;Tianhai Ji; Zhongqun Tian ; Chaoyong James Yang
Angewandte Chemie 2015 Volume 127( Issue 36) pp:10594-10599
Publication Date(Web):
DOI:10.1002/ange.201503963
Abstract
Herein, we demonstrate that a very familiar, yet underutilized, physical parameter—gas pressure—can serve as signal readout for highly sensitive bioanalysis. Integration of a catalyzed gas-generation reaction with a molecular recognition component leads to significant pressure changes, which can be measured with high sensitivity using a low-cost and portable pressure meter. This new signaling strategy opens up a new way for simple, portable, yet highly sensitive biomedical analysis in a variety of settings.
Co-reporter:Jiuxing Li, Bingqing Zhu, Zhi Zhu, Yicong Zhang, Xiujie Yao, Song Tu, Rudi Liu, Shasha Jia, and Chaoyong James Yang
Langmuir 2015 Volume 31(Issue 28) pp:7869-7876
Publication Date(Web):June 23, 2015
DOI:10.1021/acs.langmuir.5b01680
DNA conjugated gold nanorods (AuNRs) are widely applied for nanostructure assembly, gene therapy, biosensing, and drug delivery. However, it is still a great challenge to attach thiolated DNA on AuNRs, because the positively charged AuNRs readily aggregate in the presence of negatively charged DNA. This article reports an mPEG-SH/Tween 20-assisted method to load thiolated DNA on AuNRs in 1 h. Tween 20 and mPEG-SH are used to synergistically displace CTAB on the surface of AuNRs by repeated centrifugation and resuspension, and thiolated DNA are attached to AuNRs in the presence of 1 M NaCl, 100 mM MgCl2, or 100 mM citrate. AuNRs with different sizes and aspect ratios can be functionalized with DNA by this method. The number of DNA loaded on each AuNR can be easily controlled by the concentrations of mPEG-SH and Tween 20 or the ratio between DNA and AuNR. Functionalized AuNRs were used for nanoparticle assembly and cancer cell imaging to confirm that DNA anchored on the surface of AuNRs retains its hybridization and molecular recognition capability. The new method is easy, rapid, and robust for the preparation of DNA functionalized AuNRs for a variety of applications such as cancer therapy, drug delivery, self-assembly, and imaging.
Co-reporter: Zhi Zhu;Zhichao Guan;Dan Liu;Shasha Jia;Jiuxing Li;Zhichao Lei;Shuichao Lin;Tianhai Ji; Zhongqun Tian ; Chaoyong James Yang
Angewandte Chemie International Edition 2015 Volume 54( Issue 36) pp:10448-10453
Publication Date(Web):
DOI:10.1002/anie.201503963
Abstract
Herein, we demonstrate that a very familiar, yet underutilized, physical parameter—gas pressure—can serve as signal readout for highly sensitive bioanalysis. Integration of a catalyzed gas-generation reaction with a molecular recognition component leads to significant pressure changes, which can be measured with high sensitivity using a low-cost and portable pressure meter. This new signaling strategy opens up a new way for simple, portable, yet highly sensitive biomedical analysis in a variety of settings.
Co-reporter:Yuan An;Jie Wu;Bo Yang;Zhi Zhu;Mingxuan Gao
Journal of Molecular Evolution 2015 Volume 81( Issue 5-6) pp:179-185
Publication Date(Web):2015 December
DOI:10.1007/s00239-015-9703-y
Amplified in breast cancer 1 (AIB1), also known as steroid receptor coactivator 3 (SRC-3), is a transcriptional coactivator that interacts with nuclear receptors and other transcription factors to enhance their effects on target gene transcription. AIB1, which acts as a major oncogene, is highly expressed in many human cancers, and has been demonstrated to be a key regulator for tumor initiation, progression, metastasis, invasion, and survival. Recruitment of the transcriptional factor CBP/p300 by CBP/p300-interaction domain (CID) of AIB1 is essential for its transcriptional activation function. In this research, we isolated a DNA aptamer AY-3 that binds to AIB1-CID from a random oligonucleotide library using in vitro screening technology—Systematic Evolution of Ligands by EXponential enrichment (SELEX). The binding affinity of the aptamer to AIB1-CID fusion protein is in the nanomolar range. More importantly, the aptamer was found to disrupt in the interaction between p300 and AIB1. This aptamer has great potential to serve as a therapeutic agent for cancer by inhibiting the coactivation of AIB1.
Co-reporter:Jiuxing Li, Binqing Zhu, Xiujie Yao, Yicong Zhang, Zhi Zhu, Song Tu, Shasha Jia, Rudi Liu, Huaizhi Kang, and Chaoyong James Yang
ACS Applied Materials & Interfaces 2014 Volume 6(Issue 19) pp:16800
Publication Date(Web):September 4, 2014
DOI:10.1021/am504139d
Attaching thiolated DNA on gold nanoparticles (AuNPs) has been extremely important in nanobiotechnology because DNA–AuNPs combine the programmability and molecular recognition properties of the biopolymers with the optical, thermal, and catalytic properties of the inorganic nanomaterials. However, current standard protocols to attach thiolated DNA on AuNPs involve time-consuming, tedious steps and do not perform well for large AuNPs, thereby greatly restricting applications of DNA–AuNPs. Here we demonstrate a rapid and facile strategy to attach thiolated DNA on AuNPs based on the excellent stabilization effect of mPEG-SH on AuNPs. AuNPs are first protected by mPEG-SH in the presence of Tween 20, which results in excellent stability of AuNPs in high ionic strength environments and extreme pHs. A high concentration of NaCl can be applied to the mixture of DNA and AuNP directly, allowing highly efficient DNA attachment to the AuNP surface by minimizing electrostatic repulsion. The entire DNA loading process can be completed in 1.5 h with only a few simple steps. DNA-loaded AuNPs are stable for more than 2 weeks at room temperature, and they can precisely hybridize with the complementary sequence, which was applied to prepare core–satellite nanostructures. Moreover, cytotoxicity assay confirmed that the DNA–AuNPs synthesized by this method exhibit lower cytotoxicity than those prepared by current standard methods. The proposed method provides a new way to stabilize AuNPs for rapid and facile loading thiolated DNA on AuNPs and will find wide applications in many areas requiring DNA–AuNPs, including diagnosis, therapy, and imaging.Keywords: bioconjugation; DNA conjugation; Gold nanoparticle; self-assembly; surface modification; surfactants
Co-reporter:Dan Liu, Bingqian Lin, Wei Shao, Zhi Zhu, Tianhai Ji, and Chaoyong Yang
ACS Applied Materials & Interfaces 2014 Volume 6(Issue 3) pp:2131
Publication Date(Web):January 13, 2014
DOI:10.1021/am405219u
Transport of PEGylated silica nanoparticles (PSiNPs) with diameters of 100, 50, and 25 nm across the blood–brain barrier (BBB) was evaluated using an in vitro BBB model based on mouse cerebral endothelial cells (bEnd.3) cultured on transwell inserts within a chamber. In vivo animal experiments were further performed by noninvasive in vivo imaging and ex vivo optical imaging after injection via carotid artery. Confocal fluorescence studies were carried out to evaluate the uptake of PSiNPs by brain endothelial cells. The results showed that PSiNPs can traverse the BBB in vitro and in vivo. The transport efficiency of PSiNPs across BBB was found to be size-dependent, with increased particle size resulting in decreased efficiency. This work points to the potential application of small sized silica nanoparticles in brain imaging or drug delivery.Keywords: blood−brain barrier; different size; PEGylated silica nanoparticles; transportation;
Co-reporter:Guoliang Ke, Zhi Zhu, Wei Wang, Yuan Zou, Zhichao Guan, Shasha Jia, Huimin Zhang, Xuemeng Wu, and Chaoyong James Yang
ACS Applied Materials & Interfaces 2014 Volume 6(Issue 17) pp:15329
Publication Date(Web):August 11, 2014
DOI:10.1021/am503818n
Accurate sensing of the extracellular pH is a very important yet challenging task in biological and clinical applications. This paper describes the development of an amphiphilic lipid–DNA molecule as a simple yet useful cell-surface-anchored ratiometric fluorescent probe for extracellular pH sensing. The lipid–DNA probe, which consists of a hydrophobic diacyllipid tail and a hydrophilic DNA strand, is modified with two fluorescent dyes; one is pH-sensitive as pH indicator and the other is pH-insensitive as an internal reference. The lipid–DNA probe showed sensitive and reversible response to pH change in the range of 6.0–8.0, which is suitable for most extracellular studies. In addition, based on simple hydrophobic interactions with the cell membrane, the lipid–DNA probe can be easily anchored on the cell surface with negligible cytotoxicity, excellent stability, and unique ratiometric readout, thus ensuring its accurate sensing of extracellular pH. Finally, this lipid–DNA-based ratiometric pH indicator was successfully used for extracellular pH sensing of cells in 3D culture environment, demonstrating the potential applications of the sensor in biological and medical studies.Keywords: cell surface anchorage; extracellular pH sensing; lipid−DNA; ratiometric fluorescent probe
Co-reporter:Liang Cui, Zhi Zhu, Ninghang Lin, Huimin Zhang, Zhichao Guan and Chaoyong James Yang
Chemical Communications 2014 vol. 50(Issue 13) pp:1576-1578
Publication Date(Web):05 Dec 2013
DOI:10.1039/C3CC48707E
A T7 exonuclease-assisted cyclic enzymatic amplification method (CEAM) was combined with rolling circle amplification (RCA) to develop a RCA–CEAM dual amplification method for ultrasensitive detection of microRNA with excellent selectivity.
Co-reporter:Huimin Zhang, Yanling Song, Yuan Zou, Yun Ge, Yuan An, Yanli Ma, Zhi Zhu and Chaoyong James Yang
Chemical Communications 2014 vol. 50(Issue 38) pp:4891-4894
Publication Date(Web):21 Mar 2014
DOI:10.1039/C4CC01528B
A photo-reactive functional labelling reagent, diazirine phosphoramidite, was designed and synthesized for easy and flexible site-specific labelling of oligonucleotides with the diazirine moiety. The new reagent allows facile photo-crosslinking of oligonucleotide with its interacting partner for a variety of applications, including tertiary structure determination, molecular interaction study and biomarker discovery.
Co-reporter:Xiaoyan Lin, Liang Cui, Yishun Huang, Ya Lin, Yi Xie, Zhi Zhu, Bincheng Yin, Xi Chen and Chaoyong James Yang
Chemical Communications 2014 vol. 50(Issue 57) pp:7646-7648
Publication Date(Web):22 May 2014
DOI:10.1039/C4CC02184C
Based on the protective properties of carbon nanoparticles for aptamers against the digestion of nuclease, we have developed a nuclease-assisted target recycling signal amplification method for highly sensitive detection of biomolecules, such as ATP and kanamycin. The high binding specificity between aptamers and targets leads to excellent selectivity of the assay.
Co-reporter:Chunshui Lin, Zhixiong Cai, Yiru Wang, Zhi Zhu, Chaoyong James Yang, and Xi Chen
Analytical Chemistry 2014 Volume 86(Issue 14) pp:6758
Publication Date(Web):July 1, 2014
DOI:10.1021/ac501730u
A simple, rapid, label-free, and ultrasensitive fluorescence strategy for adenosine triphosphate (ATP) detection was developed using a loop DNA probe with low background noise. In this strategy, a loop DNA probe, which is the substrate for both ligation and digestion enzyme reaction, was designed. SYBR green I (SG I), a double-stranded specific dye, was applied for the readout fluorescence signal. Exonuclease I (Exo I) and exonuclease III (Exo III), sequence-independent nucleases, were selected to digest the loop DNA probe in order to minimize the background fluorescence signal. As a result, in the absence of ATP, the loop DNA was completely digested by Exo I and Exo III, leading to low background fluorescence owing to the weak electrostatic interaction between SG I and mononucleotides. On the other hand, ATP induced the ligation of the nicking site, and the sealed loop DNA resisted the digestion of Exo I and ExoIII, resulting in a remarkable increase of fluorescence response. Upon background noise reduction, the sensitivity of the ATP determination was improved significantly, and the detection limitation was found to be 1.2 pM, which is much lower than that in almost all the previously reported methods. This strategy has promise for wide application in the determination of ATP.
Co-reporter:Xilan Li, Weiyun Zhang, Lu Liu, Zhi Zhu, Gaoliang Ouyang, Yuan An, Chunyi Zhao, and Chaoyong James Yang
Analytical Chemistry 2014 Volume 86(Issue 13) pp:6596
Publication Date(Web):June 3, 2014
DOI:10.1021/ac501205q
Cancer is a major public health issue, with metastatic cancer accounting for the overwhelming majority of cancer deaths. Early diagnosis and appropriate treatment of metastatic cancer may largely prolong the survival rate and improve the quality of life for patients. In this study, we have identified a panel of DNA aptamers specifically binding to MDA-MB-231 cells derived from metastatic site-pleural effusion, with high affinity after 15 rounds of selections using the cell-based systematic evolution of ligands by exponential enrichment (SELEX) method. The selected aptamers were subjected to flow cytometry and laser confocal fluorescence microscopy to evaluate their binding affinity and selectivity. The aptamer LXL-1 with the highest abundance in the enriched library demonstrated a low Kd value and excellent selectivity for the recognition of the metastatic breast cancer cells. Tissue imaging results showed that truncated aptamer sequence LXL-1-A was highly specific to the corresponding tumor tissue and displayed 76% detection rate against breast cancer tissue with metastasis in regional lymph nodes. Therefore, on the basis of its excellent targeting properties and functional versatility, LXL-1-A holds great potential to be used as a molecular imaging probe for the detection of breast cancer metastasis. Our result clearly demonstrates that metastatic-cell-based SELEX can be used to generate DNA ligands specifically recognizing metastatic cancer cells, which is of great significance for metastatic cancer diagnosis and treatment.
Co-reporter:Zhi Zhu, Yanling Song, Cong Li, Yuan Zou, Ling Zhu, Yuan An, and Chaoyong James Yang
Analytical Chemistry 2014 Volume 86(Issue 12) pp:5881
Publication Date(Web):May 27, 2014
DOI:10.1021/ac501423g
A novel method, monoclonal surface display SELEX (MSD-SELEX), has been designed for simple, rapid, efficient, and cost-effective enrichment and identification of aptamers from a library of monoclonal DNA-displaying beads produced via highly parallel single-molecule emulsion PCR. The approach was successfully applied for the identification of high-affinity aptamers that bind specifically to different types of targets, including cancer biomarker protein EpCAM and small toxin molecule aflatoxin B1. Compared to the conventional sequencing-chemical synthesis-screening work flow, MSD-SELEX avoids large-scale DNA sequencing, expensive and time-consuming DNA synthesis, and labor-intensive screening of large populations of candidates, thus offering a new approach for simple, rapid, efficient, and cost-effective aptamer identification for a wide variety of applications.
Co-reporter:Zhichao Guan, Shasha Jia, Zhi Zhu, Mingxia Zhang, and Chaoyong James Yang
Analytical Chemistry 2014 Volume 86(Issue 5) pp:2789
Publication Date(Web):February 10, 2014
DOI:10.1021/ac500088m
Microfabricated devices are suitable for single-cell analysis due to their high throughput, compatible dimensions and controllable microenvironment. However, existing devices for single-cell culture and analysis encounter some limitations, such as nutrient depletion, random cell migration and complicated fluid shear influence. Moreover, most of the single-cell culture and analysis devices are based on 2D cell culture conditions, even though 3D cell culture methods have been demonstrated to better mimic the real cell microenvironment in vivo. To solve these problems, herein we develop a microcollagen gel array (μCGA) based approach for high-throughput long-term single-cell culture and single-cell analysis under 3D culture conditions. Type-I collagen, a well-established 3D cell culture medium, was used as the scaffold for 3D cell growth. A 2 × 2 cm PDMS chip with 10 000 μCGA units was fabricated to encapsulate thousands of single cells in less than 15 min. Single cells were able to be confined and survive in μCGA units for more than 1 month. The capability of large-scale and long-term single-cell 3D culture under open culture conditions allows us to study cellular proliferation heterogeneity and drug cytotoxicity at the single-cell level. Compared with existing devices for single-cell analysis, μCGA solves the problems of nutrient depletion and random cellular migration, avoids the influence of complicated fluid shear, and mimics the real 3D growth environment in vivo, thereby providing a feasible 3D long-term single-cell culture method for single-cell analysis and drug screening.
Co-reporter:Yishun Huang, Yanli Ma, Yahong Chen, Xuemeng Wu, Luting Fang, Zhi Zhu, and Chaoyong James Yang
Analytical Chemistry 2014 Volume 86(Issue 22) pp:11434
Publication Date(Web):October 23, 2014
DOI:10.1021/ac503540q
Because of the severe health risks associated with lead pollution, rapid, sensitive, and portable detection of low levels of Pb2+ in biological and environmental samples is of great importance. In this work, a Pb2+-responsive hydrogel was prepared using a DNAzyme and its substrate as cross-linker for rapid, sensitive, portable, and quantitative detection of Pb2+. Gold nanoparticles (AuNPs) were first encapsulated in the hydrogel as an indicator for colorimetric analysis. In the absence of lead, the DNAzyme is inactive, and the substrate cross-linker maintains the hydrogel in the gel form. In contrast, the presence of lead activates the DNAzyme to cleave the substrate, decreasing the cross-linking density of the hydrogel and resulting in dissolution of the hydrogel and release of AuNPs for visual detection. As low as 10 nM Pb2+ can be detected by the naked eye. Furthermore, to realize quantitative visual detection, a volumetric bar-chart chip (V-chip) was used for quantitative readout of the hydrogel system by replacing AuNPs with gold–platinum core–shell nanoparticles (Au@PtNPs). The Au@PtNPs released from the hydrogel upon target activation can efficiently catalyze the decomposition of H2O2 to generate a large volume of O2. The gas pressure moves an ink bar in the V-chip for portable visual quantitative detection of lead with a detection limit less than 5 nM. The device was able to detect lead in digested blood with excellent accuracy. The method developed can be used for portable lead quantitation in many applications. Furthermore, the method can be further extended to portable visual quantitative detection of a variety of targets by replacing the lead-responsive DNAzyme with other DNAzymes.
Co-reporter:Jiangwei Tian;Dr. Lin Ding; Huangxian Ju;Dr. Yongchao Yang;Xilan Li; Zhen Shen;Dr. Zhi Zhu; Jun-Sheng Yu; Chaoyong James Yang
Angewandte Chemie International Edition 2014 Volume 53( Issue 36) pp:9544-9549
Publication Date(Web):
DOI:10.1002/anie.201405490
Abstract
Simultaneous targeted cancer imaging, therapy and real-time therapeutic monitoring can prevent over- or undertreatment. This work describes the design of a multifunctional nanomicelle for recognition and precise near-infrared (NIR) cancer therapy. The nanomicelle encapsulates a new pH-activatable fluorescent probe and a robust NIR photosensitizer, R16FP, and is functionalized with a newly screened cancer-specific aptamer for targeting viable cancer cells. The fluorescent probe can light up the lysosomes for real-time imaging. Upon NIR irradiation, R16FP-mediated generation of reactive oxygen species causes lysosomal destruction and subsequently trigger lysosomal cell death. Meanwhile the fluorescent probe can reflect the cellular status and in situ visualize the treatment process. This protocol can provide molecular information for precise therapy and therapeutic monitoring.
Co-reporter: Zhi Zhu;Zhichao Guan;Shasha Jia;Zhichao Lei;Dr. Shuichao Lin;Huimin Zhang;Yanli Ma; Zhong-Qun Tian ; Chaoyong James Yang
Angewandte Chemie International Edition 2014 Volume 53( Issue 46) pp:12503-12507
Publication Date(Web):
DOI:10.1002/anie.201405995
Abstract
Point-of-care testing (POCT) with the advantages of speed, simplicity, portability, and low cost is critical for the measurement of analytes in a variety of environments where access to laboratory infrastructure is lacking. While qualitative POCTs are widely available, quantitative POCTs present significant challenges. Here we describe a novel method that integrates an Au core/Pt shell nanoparticle (Au@PtNP) encapsulated target-responsive hydrogel with a volumetric bar-chart chip (V-Chip) for quantitative POCT. Upon target introduction, the hydrogel immediately dissolves and releases Au@PtNPs, which can efficiently catalyze the decomposition of H2O2 to generate a large volume of O2 to move of an ink bar in the V-Chip. The concentration of the target introduced can be visually quantified by reading the traveling distance of the ink bar. This method has the potential to be used for portable and quantitative detection of a wide range of targets without any external instrument.
Co-reporter: Zhi Zhu;Zhichao Guan;Shasha Jia;Zhichao Lei;Dr. Shuichao Lin;Huimin Zhang;Yanli Ma; Zhong-Qun Tian ; Chaoyong James Yang
Angewandte Chemie 2014 Volume 126( Issue 46) pp:12711-12715
Publication Date(Web):
DOI:10.1002/ange.201405995
Abstract
Point-of-care testing (POCT) with the advantages of speed, simplicity, portability, and low cost is critical for the measurement of analytes in a variety of environments where access to laboratory infrastructure is lacking. While qualitative POCTs are widely available, quantitative POCTs present significant challenges. Here we describe a novel method that integrates an Au core/Pt shell nanoparticle (Au@PtNP) encapsulated target-responsive hydrogel with a volumetric bar-chart chip (V-Chip) for quantitative POCT. Upon target introduction, the hydrogel immediately dissolves and releases Au@PtNPs, which can efficiently catalyze the decomposition of H2O2 to generate a large volume of O2 to move of an ink bar in the V-Chip. The concentration of the target introduced can be visually quantified by reading the traveling distance of the ink bar. This method has the potential to be used for portable and quantitative detection of a wide range of targets without any external instrument.
Co-reporter:Ling Yan ; Zhi Zhu ; Yuan Zou ; Yishun Huang ; Dewen Liu ; Shasha Jia ; Dunming Xu ; Min Wu ; Yu Zhou ; Shuang Zhou
Journal of the American Chemical Society 2013 Volume 135(Issue 10) pp:3748-3751
Publication Date(Web):January 22, 2013
DOI:10.1021/ja3114714
Portable devices with the advantages of rapid, on-site, user-friendly, and cost-effective assessment are widely applied in daily life. However, only a limited number of quantitative portable devices are commercially available, among which the personal glucose meter (PGM) is the most successful example and has been the most widely used. However, PGMs can detect only blood glucose as the unique target. Here we describe a novel design that combines a glucoamylase-trapped aptamer-cross-linked hydrogel with a PGM for portable and quantitative detection of non-glucose targets. Upon target introduction, the hydrogel collapses to release glucoamylase, which catalyzes the hydrolysis of amylose to produce a large amount of glucose for quantitative readout by the PGM. With the advantages of low cost, rapidity, portability, and ease of use, the method reported here has the potential to be used by the public for portable and quantitative detection of a wide range of non-glucose targets.
Co-reporter:Yuan Zou, Jie Chen, Zhi Zhu, Lianyu Lu, Yishun Huang, Yanling Song, Huimin Zhang, Huaizhi Kang and Chaoyong James Yang
Chemical Communications 2013 vol. 49(Issue 77) pp:8716-8718
Publication Date(Web):29 Jul 2013
DOI:10.1039/C3CC44188A
A pair of single-molecule photo-responsive DNA nanoscissors for DNA cleavage based on the regulation of substrate binding affinity was designed and fabricated. Compared with other DNA nanomachines, our DNA nanoscissors have the advantages of a clean switching mechanism, as well as robust and highly reversible operation.
Co-reporter:Xiaoyan Lin, Cheng Zhang, Yishun Huang, Zhi Zhu, Xi Chen and Chaoyong James Yang
Chemical Communications 2013 vol. 49(Issue 65) pp:7243-7245
Publication Date(Web):18 Jun 2013
DOI:10.1039/C3CC43224F
Based on backbone-modified molecular beacons and duplex-specific nuclease, we have developed a target recycling amplification method for highly sensitive and selective miRNA detection. The combination of a low fluorescence background of 2-OMe-RNA modified MB and nuclease-assisted signal amplification leads to ultrahigh assay sensitivity, and the powerful discriminating ability of MB enables the differentiation of highly similar miRNAs with one-base difference, both of which are of great significance to miRNA detection.
Co-reporter:Yanling Song, Zhi Zhu, Yuan An, Weiting Zhang, Huimin Zhang, Dan Liu, Chundong Yu, Wei Duan, and Chaoyong James Yang
Analytical Chemistry 2013 Volume 85(Issue 8) pp:4141
Publication Date(Web):March 12, 2013
DOI:10.1021/ac400366b
Epithelial cell adhesion molecule (EpCAM) is overexpressed in most solid cancers and is an ideal antigen for clinical applications in cancer diagnosis, prognosis, imaging, and therapy. Currently, most of the EpCAM-based diagnostic, prognostic, and therapeutic strategies rely on the anti-EpCAM antibody. However, the use of EpCAM antibody is restricted due to its large size and instability. In this study, we have successfully identified DNA aptamers that selectively bind human recombinant EpCAM protein. The aptamers can specifically recognize a number of live human cancer cells derived from breast, colorectal, and gastric cancers that express EpCAM but not bind to EpCAM-negative cells. Among the aptamer sequences identified, a hairpin-structured sequence SYL3 was optimized in length, resulting in aptamer sequence SYL3C. The Kd values of the SYL3C aptamer against breast cancer cell line MDA-MB-231 and gastric cancer cell line Kato III were found to be 38 ± 9 and 67 ± 8 nM, respectively, which are better than that of the full-length SYL3 aptamer. Flow cytometry analysis results indicated that the SYL3C aptamer was able to recognize target cancer cells from mixed cells in cell media. When used to capture cancer cells, up to 63% cancer cell capture efficiency was achieved with about 80% purity. With the advantages of small size, easy synthesis, good stability, high binding affinity, and selectivity, the DNA aptamers reported here against cancer biomarker EpCAM will facilitate the development of novel targeted cancer therapy, cancer cell imaging, and circulating tumor cell detection.
Co-reporter:Liang Cui, Zirong Chen, Zhi Zhu, Xiaoyan Lin, Xi Chen, and Chaoyong James Yang
Analytical Chemistry 2013 Volume 85(Issue 4) pp:2269
Publication Date(Web):January 16, 2013
DOI:10.1021/ac303179z
RNA probes constitute an important class of functional nucleic acids (FNAs). However, because of their notorious vulnerability to enzymatic degradation, extremely careful and special protocols must be followed when dealing with RNA probes. To fully use the large number of RNA FNAs available for bioanalysis and biomedicine, it is important to explore effective methods to protect RNA probes from enzymatic digestion. In this work, we systematically demonstrate that graphene oxide (GO) can effectively protect RNA probes from enzymatic digestion. Based on this finding, we propose an effective way to design robust RNA biosensors by simply mixing RNA probes with GO for analysis of nucleic acids, proteins, and small molecules. The entire assay is sensitive, selective, rapid, and more importantly, does not require any special protocols. The ability to protect ssRNA from enzymatic digestion by GO offers an exciting new way to stabilize ssRNA, which will not only provide new opportunities to utilize the large number of currently available, yet rarely explored, RNA FNAs for bioanalysis but also offer a new solution to protect important ssRNA molecules, such as microRNA and antisense ssRNA, for a great variety of biomedical applications.
Co-reporter:Xiangling Xiong;Dr. Haipeng Liu;Dr. Zilong Zhao;Dr. Meghan B. Altman;Dr. Dalia Lopez-Colon; Chaoyong James Yang; Lung-Ji Chang; Chen Liu; Weihong Tan
Angewandte Chemie International Edition 2013 Volume 52( Issue 5) pp:1472-1476
Publication Date(Web):
DOI:10.1002/anie.201207063
Co-reporter:Liang Cui;Yanling Song;Guoliang Ke;Zhichao Guan;Huimin Zhang;Ya Lin;Yishun Huang;Dr. Zhi Zhu; Chaoyong James Yang
Chemistry - A European Journal 2013 Volume 19( Issue 32) pp:10442-10451
Publication Date(Web):
DOI:10.1002/chem.201301292
Abstract
Recently, the binding ability of DNA on GO and resulting nuclease resistance have attracted increasing attention, leading to new applications both in vivo and in vitro. In vivo, nucleic acids absorbed on GO can be effectively protected from enzymatic degradation and biological interference in complicated samples, making it useful for targeted delivery, gene regulation, intracellular detection and imaging with high uptake efficiencies, high intracellular stability, and very low toxicity. In vitro, the adsorption of ssDNA on GO surface and desorption of dsDNA or well-folded ssDNA from GO surface result in the protection and deprotection of DNA from nucleic digestion, respectively, which has led to target-triggered cyclic enzymatic amplification methods (CEAM) for amplified detection of analytes with sensitivity 2–3 orders of magnitude higher than that of 1:1 binding strategies. This Concept article explores some of the latest developments in this field.
Co-reporter:Xiangling Xiong;Dr. Haipeng Liu;Dr. Zilong Zhao;Dr. Meghan B. Altman;Dr. Dalia Lopez-Colon; Chaoyong James Yang; Lung-Ji Chang; Chen Liu; Weihong Tan
Angewandte Chemie 2013 Volume 125( Issue 5) pp:1512-1516
Publication Date(Web):
DOI:10.1002/ange.201207063
Co-reporter:Guoliang Ke ; Chunming Wang ; Yun Ge ; Nanfeng Zheng ; Zhi Zhu
Journal of the American Chemical Society 2012 Volume 134(Issue 46) pp:18908-18911
Publication Date(Web):November 5, 2012
DOI:10.1021/ja3082439
Noninvasive and accurate measurement of intracellular temperature is of great significance in biology and medicine. This paper describes a safe, stable, and accurate intracellular nano-thermometer based on an l-DNA molecular beacon (l-MB), a dual-labeled hairpin oligonucleotide built from the optical isomer of naturally occurring d-DNA. Relying on the temperature-responsive hairpin structure and the FRET signaling mechanism of MBs, the fluorescence of l-MBs is quenched below the melting temperature and enhanced with increasing temperature. Because of the excellent reversibility and tunable response range, l-MBs are very suitable for temperature sensing. More importantly, the non-natural l-DNA backbone prevents the l-MBs from binding to cellular nucleic acids and proteins as well as from being digested by nucleases inside the cells, thus ensuring excellent stability and accuracy of the nano-thermometer in a complex cellular environment. The l-MB nano-thermometer was used for the photothermal study of Pd nanosheets in living cells, establishing the nano-thermometer as a useful tool for intracellular temperature measurement.
Co-reporter:Liang Cui, Xiaoyan Lin, Ninghang Lin, Yanling Song, Zhi Zhu, Xi Chen and Chaoyong James Yang
Chemical Communications 2012 vol. 48(Issue 2) pp:194-196
Publication Date(Web):05 Oct 2011
DOI:10.1039/C1CC15412E
Based on graphene oxide-protected DNA probes, we have developed a cyclic enzymatic amplification method for sensitive miRNA detection in complex biological samples. By using the quenching nature of graphene oxide for multiple fluorophores, this method can distinguish highly similar miRNA sequences and detect them simultaneously.
Co-reporter:Yanling Song, Weiting Zhang, Yuan An, Liang Cui, Chundong Yu, Zhi Zhu and Chaoyong James Yang
Chemical Communications 2012 vol. 48(Issue 4) pp:576-578
Publication Date(Web):07 Nov 2011
DOI:10.1039/C1CC15777A
We have combined an allosteric molecular beacon for target recognition and guanine-rich DNAzyme for signal amplification to develop a new platform for visual detection of nucleic acids with single-base mismatch detection capability. The fully DNA-structured platform can undergo color change in response to target DNA/RNA, which enables sensitive and selective visual detection in biological samples.
Co-reporter:Cong Li, Ling Zhu, Zhi Zhu, Hao Fu, Gareth Jenkins, Chunming Wang, Yuan Zou, Xin Lu and Chaoyong James Yang
Chemical Communications 2012 vol. 48(Issue 67) pp:8347-8349
Publication Date(Web):29 Jun 2012
DOI:10.1039/C2CC32919K
In this work we studied how backbone chemical modifications, such as 2′-O-methyl, phosphorothioate, L-form nucleotides and locked nucleic acid, on G-quadruplex based DNAzymes would affect their peroxidase activity. Our results indicate that 2′-O-methyl modification facilitates the formation of a perfectly compacted parallel structure and significantly promotes peroxidase activity of G-quadruplex based DNAzymes.
Co-reporter:Zhi Zhu, Wenhua Zhang, Xuefei Leng, Mingxia Zhang, Zhichao Guan, Jiangquan Lu and Chaoyong James Yang
Lab on a Chip 2012 vol. 12(Issue 20) pp:3907-3913
Publication Date(Web):18 Jun 2012
DOI:10.1039/C2LC40461C
Genetic alternations can serve as highly specific biomarkers to distinguish fatal bacteria or cancer cells from their normal counterparts. However, these mutations normally exist in very rare amount in the presence of a large excess of non-mutated analogs. Taking the notorious pathogen E. coli O157:H7 as the target analyte, we have developed an agarose droplet-based microfluidic ePCR method for highly sensitive, specific and quantitative detection of rare pathogens in the high background of normal bacteria. Massively parallel singleplex and multiplex PCR at the single-cell level in agarose droplets have been successfully established. Moreover, we challenged the system with rare pathogen detection and realized the sensitive and quantitative analysis of a single E. coli O157:H7 cell in the high background of 100000 excess normal K12 cells. For the first time, we demonstrated rare pathogen detection through agarose droplet microfluidic ePCR. Such a multiplex single-cell agarose droplet amplification method enables ultra-high throughput and multi-parameter genetic analysis of large population of cells at the single-cell level to uncover the stochastic variations in biological systems.
Co-reporter:Liang Cui, Yuan Zou, Ninghang Lin, Zhi Zhu, Gareth Jenkins, and Chaoyong James Yang
Analytical Chemistry 2012 Volume 84(Issue 13) pp:5535
Publication Date(Web):June 11, 2012
DOI:10.1021/ac300182w
Fluorescence anisotropy (FA) is a reliable and excellent choice for fluorescence sensing. One of the key factors influencing the FA value for any molecule is the molar mass of the molecule being measured. As a result, the FA method with functional nucleic acid aptamers has been limited to macromolecules such as proteins and is generally not applicable for the analysis of small molecules because their molecular masses are relatively too small to produce observable FA value changes. We report here a molecular mass amplifying strategy to construct anisotropy aptamer probes for small molecules. The probe is designed in such a way that only when a target molecule binds to the probe does it activate its binding ability to an anisotropy amplifier (a high molecular mass molecule such as protein), thus significantly increasing the molecular mass and FA value of the probe/target complex. Specifically, a mass amplifying probe (MAP) consists of a targeting aptamer domain against a target molecule and molecular mass amplifying aptamer domain for the amplifier protein. The probe is initially rendered inactive by a small blocking strand partially complementary to both target aptamer and amplifier protein aptamer so that the mass amplifying aptamer domain would not bind to the amplifier protein unless the probe has been activated by the target. In this way, we prepared two probes that constitute a target (ATP and cocaine respectively) aptamer, a thrombin (as the mass amplifier) aptamer, and a fluorophore. Both probes worked well against their corresponding small molecule targets, and the detection limits for ATP and cocaine were 0.5 μM and 0.8 μM, respectively. More importantly, because FA is less affected by environmental interferences, ATP in cell media and cocaine in urine were directly detected without any tedious sample pretreatment. Our results established that our molecular mass amplifying strategy can be used to design aptamer probes for rapid, sensitive, and selective detection of small molecules by means of FA in complex biological samples.
Co-reporter:Huifa Zhang, Gareth Jenkins, Yuan Zou, Zhi Zhu, and Chaoyong James Yang
Analytical Chemistry 2012 Volume 84(Issue 8) pp:3599
Publication Date(Web):March 27, 2012
DOI:10.1021/ac2033084
A microfluidic device for performing single copy, emulsion Reverse Transcription Polymerase Chain Reaction (RT-PCR) within agarose droplets is presented. A two-aqueous-inlet emulsion droplet generator was designed and fabricated to produce highly uniform monodisperse picoliter agarose emulsion droplets with RT-PCR reagents in carrier oil. Template RNA or cells were delivered from one inlet with RT-PCR reagents/cell lysis buffer delivered separately from the other. Efficient RNA/cell encapsulation and RT-PCR at the single copy level was achieved in agarose-in-oil droplets, which, after amplification, can be solidified into agarose beads for further analysis. A simple and efficient method to graft primer to the polymer matrix using 5′-acrydite primer was developed to ensure highly efficient trapping of RT-PCR products in agarose. High-throughput single RNA molecule/cell RT-PCR was demonstrated in stochastically diluted solutions. Our results indicate that single-molecule RT-PCR can be efficiently carried out in agarose matrix. Single-cell RT-PCR was successfully performed which showed a clear difference in gene expression level of EpCAM, a cancer biomarker gene, at the single-cell level between different types of cancer cells. This work clearly demonstrates for the first time, single-copy RT-PCR in agarose droplets. We believe this will open up new possibilities for viral RNA detection and single-cell transcription analysis.
Co-reporter:Wei Yun Zhang, Wenhua Zhang, Zhiyuan Liu, Cong Li, Zhi Zhu, and Chaoyong James Yang
Analytical Chemistry 2012 Volume 84(Issue 1) pp:350
Publication Date(Web):November 21, 2011
DOI:10.1021/ac2026942
We have developed a novel method for efficiently screening affinity ligands (aptamers) from a complex single-stranded DNA (ssDNA) library by employing single-molecule emulsion polymerase chain reaction (PCR) based on the agarose droplet microfluidic technology. In a typical systematic evolution of ligands by exponential enrichment (SELEX) process, the enriched library is sequenced first, and tens to hundreds of aptamer candidates are analyzed via a bioinformatic approach. Possible candidates are then chemically synthesized, and their binding affinities are measured individually. Such a process is time-consuming, labor-intensive, inefficient, and expensive. To address these problems, we have developed a highly efficient single-molecule approach for aptamer screening using our agarose droplet microfluidic technology. Statistically diluted ssDNA of the pre-enriched library evolved through conventional SELEX against cancer biomarker Shp2 protein was encapsulated into individual uniform agarose droplets for droplet PCR to generate clonal agarose beads. The binding capacity of amplified ssDNA from each clonal bead was then screened via high-throughput fluorescence cytometry. DNA clones with high binding capacity and low Kd were chosen as the aptamer and can be directly used for downstream biomedical applications. We have identified an ssDNA aptamer that selectively recognizes Shp2 with a Kd of 24.9 nM. Compared to a conventional sequencing–chemical synthesis–screening work flow, our approach avoids large-scale DNA sequencing and expensive, time-consuming DNA synthesis of large populations of DNA candidates. The agarose droplet microfluidic approach is thus highly efficient and cost-effective for molecular evolution approaches and will find wide application in molecular evolution technologies, including mRNA display, phage display, and so on.
Co-reporter:Ling Zhu, Cong Li, Zhi Zhu, Dewen Liu, Yuan Zou, Chunming Wang, Hao Fu, and Chaoyong James Yang
Analytical Chemistry 2012 Volume 84(Issue 19) pp:8383
Publication Date(Web):September 7, 2012
DOI:10.1021/ac301899h
Because of their ability to greatly enhance the low natural peroxidase activity of hemin, G-quadruplex-based DNAzymes have been widely used as an alternative to peroxidases for many colorimetric, chemiluminescent, or visual detections of metal ions, small molecules, nucleic acids, proteins, and cancer cells. To obtain G-quadruplex-based DNAzymes with better peroxidase activity, we designed three 81-nt ssDNA libraries containing 25%, 35%, and 45% guanine bases, respectively, at the 45-nt random regions to evolve hemin-binding DNA aptamers using hemin–agarose beads by SELEX (systematic evolution of ligands by exponential enrichment). Some G-rich sequences were obtained after 6 rounds of selection and optimized for stronger binding affinity to hemin and higher peroxidase activity. Our results show that the truncated aptamer [B7]-3-0 folds into compact parallel G-quadruplex structure and exhibits the highest peroxidase activity and strong binding affinity to hemin with 29 ± 4 nM of Kd. It was found that the core G-motifs sequences with 5′-flanking nucleotides exhibit higher peroxidase activity than those with 3′-flanking nucleotides. The numbers of 5′-flanking nucleotides also influence peroxidase activity. In addition, 2′-O-methyl modification facilitates the self-assembly of parallel G-quadruplex [B7]-3-0 and significantly promotes peroxidase activity. This study identifies a G-quadruplex sequence with peroxidase-like activity higher than any other sequences reported so far, which could be potentially used to improve the analytical performance of a wide variety of peroxidase-based bioassays.
Co-reporter:Zhi Zhu;Gareth Jenkins;Wenhua Zhang
Analytical and Bioanalytical Chemistry 2012 Volume 403( Issue 8) pp:2127-2143
Publication Date(Web):2012 June
DOI:10.1007/s00216-012-5914-x
The application of microfluidic droplet PCR for single-molecule amplification and analysis has recently been extensively studied. Microfluidic droplet technology has the advantages of compartmentalizing reactions into discrete volumes, performing highly parallel reactions in monodisperse droplets, reducing cross-contamination between droplets, eliminating PCR bias and nonspecific amplification, as well as enabling fast amplification with rapid thermocycling. Here, we have reviewed the important technical breakthroughs of microfluidic droplet PCR in the past five years and their applications to single-molecule amplification and analysis, such as high-throughput screening, next generation DNA sequencing, and quantitative detection of rare mutations. Although the utilization of microfluidic droplet single-molecule PCR is still in the early stages, its great potential has already been demonstrated and will provide novel solutions to today’s biomedical engineering challenges in single-molecule amplification and analysis.
Co-reporter:Xue-Qin Zhao, Jie Wu, Jing-Hong Liang, Jia-Wei Yan, Zhi Zhu, Chaoyong James Yang, and Bing-Wei Mao
The Journal of Physical Chemistry B 2012 Volume 116(Issue 37) pp:11397-11404
Publication Date(Web):August 28, 2012
DOI:10.1021/jp303518b
With widespread applications in biosensors, diagnostics, and therapeutics, much investigation has been made in the structure of the G-quadruplexes and mechanism of their interactions with protein targets. However, in view of AFM based single-molecule force spectroscopic (SMFS) studies of G-quadruplex systems, only bimolecular approaches have been employed. In this article, we present an improved dual-labeling approach for surface immobilization of G-quadruplex DNA apatmers for investigation of intramolecular interaction from an integral unimolecular G-quadruplex system. The melting force of HJ24 G-quadruplex aptamer in the presence of K+ has been successfully measured. It has been found that dynamic equilibrium exists between unfolding and folding structures of the HJ24 aptamer even in pure water. We also investigated the interactions between the HJ24 aptamer and its target protein (Shp2) under the same solution condition. The HJ24/Shp2 unbinding force in the absence of K+, 42.0 pN, is about 50% smaller than that in the presence of K+, 61.7 pN. The great reduction in force in the absence of K+ suggests that the stability of G-quadruplex secondary structure is important for a stable HJ24/Shp2 binding. The methodology developed and demonstrated in this work is applicable for studying the stability of secondary structures of other unimolecular G-quadruplex aptamers and their interactions with target proteins.
Co-reporter:Chunming Wang, Zhi Zhu, Yanling Song, Hui Lin, Chaoyong James Yang and Weihong Tan
Chemical Communications 2011 vol. 47(Issue 20) pp:5708-5710
Publication Date(Web):18 Apr 2011
DOI:10.1039/C1CC10481K
We have constructed a novel class of light-activatable caged molecular beacons (cMBs) that are caged by locking two stems with a photo-labile biomolecular interaction or covalent bond. With the cMBs, the nucleic acid hybridization process can be easily controlled with light, which offers the possibility for a high spatiotemporal resolution study of intracellular mRNAs.
Co-reporter:Haoxue Lin, Yuan Zou, Yishun Huang, Jie Chen, Wei Yun Zhang, Zhixia Zhuang, Gareth Jenkins and Chaoyong James Yang
Chemical Communications 2011 vol. 47(Issue 33) pp:9312-9314
Publication Date(Web):31 May 2011
DOI:10.1039/C1CC12290H
We propose the use of DNAzyme as a crosslinker of hydrogel to develop a catalytic platform for the sensing of metal ions. The DNAzyme crosslinked hydrogel can undergo gel–sol transition in response to Cu2+ ions, which enables sensitive visual detection of Cu2+ by observing the release of pre-trapped AuNPs.
Co-reporter:Jia Hu;Jie Wu;Cong Li;Ling Zhu;Dr. Wei Yun Zhang;Guiping Kong; Zhongxian Lu; Chaoyong James Yang
ChemBioChem 2011 Volume 12( Issue 3) pp:424-430
Publication Date(Web):
DOI:10.1002/cbic.201000470
Abstract
Shp2 is a member of the protein tyrosine phosphatase (PTP) family, which regulates a variety of cellular processes including cell growth, differentiation, mitotic cycle, and oncogenic transformation. Using a recombinant Shp2-GST protein as the target and GST as a counter target, we have identified two classes of single-stranded DNA aptamers that selectively bind to Shp2 with a Kd in the nanomolar range. Structural studies of the most abundant sequence in the enriched library, HJ24, revealed a parallel G-quadruplex as the core binding domain. Furthermore, this aptamer was found to be an effective inhibitor of Shp2 phosphatase, an effect which was readily reversed by using the cDNA of HJ24. In view of these characteristics, this aptamer has the potential to be used for further development of Shp2 assays and therapeutics for the treatment of Shp2-dependent cancers and other diseases.
Co-reporter:Yanling Song;Liang Cui;Jie Wu;Weiting Zhang;Dr. Wei Yun Zhang;Dr. Huaizhi Kang ; Chaoyong James Yang
Chemistry - A European Journal 2011 Volume 17( Issue 33) pp:
Publication Date(Web):
DOI:10.1002/chem.201190165
Co-reporter:Yanling Song;Liang Cui;Jie Wu;Weiting Zhang;Dr. Wei Yun Zhang;Dr. Huaizhi Kang ; Chaoyong James Yang
Chemistry - A European Journal 2011 Volume 17( Issue 33) pp:9042-9046
Publication Date(Web):
DOI:10.1002/chem.201101353
Co-reporter:Dr. Kelong Wang;Mingxu You;Dr. Yan Chen;Da Han;Zhi Zhu;Jin Huang;Dr. Kathryn Williams; Chaoyong James Yang;Dr. Weihong Tan
Angewandte Chemie 2011 Volume 123( Issue 27) pp:6222-6225
Publication Date(Web):
DOI:10.1002/ange.201008053
Co-reporter:Jin Huang;Dr. Yanrong Wu;Dr. Yan Chen;Zhi Zhu; Xiaohai Yang; Chaoyong James Yang; Kemin Wang; Weihong Tan
Angewandte Chemie 2011 Volume 123( Issue 2) pp:421-424
Publication Date(Web):
DOI:10.1002/ange.201005375
Co-reporter:Dr. Kelong Wang;Mingxu You;Dr. Yan Chen;Da Han;Zhi Zhu;Jin Huang;Dr. Kathryn Williams; Chaoyong James Yang;Dr. Weihong Tan
Angewandte Chemie International Edition 2011 Volume 50( Issue 27) pp:6098-6101
Publication Date(Web):
DOI:10.1002/anie.201008053
Co-reporter:Jin Huang;Dr. Yanrong Wu;Dr. Yan Chen;Zhi Zhu; Xiaohai Yang; Chaoyong James Yang; Kemin Wang; Weihong Tan
Angewandte Chemie International Edition 2011 Volume 50( Issue 2) pp:401-404
Publication Date(Web):
DOI:10.1002/anie.201005375
Co-reporter:Chaoyong James Yang, Liang Cui, Jiahao Huang, Ling Yan, Xiaoyan Lin, Chunming Wang, Wei Yun Zhang, Huaizhi Kang
Biosensors and Bioelectronics 2011 Volume 27(Issue 1) pp:119-124
Publication Date(Web):15 September 2011
DOI:10.1016/j.bios.2011.06.027
Sensitive analysis or monitoring of biomolecules and small molecules is very important for many biological researches, clinical diagnosis and forensic investigations. As a sequence-independent exonuclease, Exonuclease III (Exo III) has been widely used for amplified detection of proteins and nucleic acids where displacing probes or molecular beacons are used as the signaling probes. However, displacing probes suffer slow hybridization rate and high background signal and molecular beacons are difficult to design and prone to undesired nonspecific interactions. Herein, we report a new type of probes called linear molecular beacons (LMBs) for use in Exo III amplification assays to improve hybridization kinetics and reduce background noises. LMBs are linear oligonucleotide probes with a fluorophore and quencher attached to 3′ terminal and penultimate nucleotides, respectively. Compared to conventional molecular beacons and displacing probes, LMBs are easy to design and synthesize. More importantly, LMBs have a much lower background noise and allow faster reaction rates. Using LMBs in cyclic Exo III amplification assay, ultrasensitive nucleic acid detection methods were developed with a detection limit of less than 120 fM, which is 2 orders of magnitude lower than that of conventional molecular beacons or displacing probes-based Exo III amplification assays. Furthermore, LMBs can be extended as universal probes for detection of non-nucleic acid molecules such as cocaine with high sensitivity. These results demonstrate that the combination of Exo III amplification and LMB signaling provides a general method for ultrasensitive and selective detection of a wide range of targets.Highlights• A linear molecular beacon (LMB) for use with Exo III has been developed. • LMB is a linear oligonucleotide probe with a fluorophore and quencher attached to 3′terminal and penultimate nucleotides respectively. • LMB does not require any intramolecular structure and thus is very easy to design. • LMB offers higher signal-to-background ratio and faster hybridization kinetics. • LMB can be used with Exo III for rapid, selective and ultrahigh sensitive detection of nucleic acid and non-nucleic acid targets.
Co-reporter:Xuefei Leng, Wenhua Zhang, Chunming Wang, Liang Cui and Chaoyong James Yang
Lab on a Chip 2010 vol. 10(Issue 21) pp:2841-2843
Publication Date(Web):13 Sep 2010
DOI:10.1039/C0LC00145G
An agarose droplet method was developed for highly parallel and efficient single molecule emulsion PCR. The method capitalizes on the unique thermoresponsive sol–gel switching property of agarose for highly efficient DNA amplification and amplicon trapping. Uniform agarose solution droplets generated via a microfluidic chip serve as robust and inert nanolitre PCR reactors for single copy DNA molecule amplification. After PCR, agarose droplets are gelated to form agarose beads, trapping all amplicons in each reactor to maintain the monoclonality of each droplet. This method does not require cocapsulation of primer labeled microbeads, allows high throughput generation of uniform droplets and enables high PCR efficiency, making it a promising platform for many single copy genetic studies.
Co-reporter:Liang Cui, Guoliang Ke, Chunming Wang and Chaoyong James Yang
Analyst 2010 vol. 135(Issue 8) pp:2069-2073
Publication Date(Web):18 Jun 2010
DOI:10.1039/C0AN00215A
Based on Exonuclease III (Exo III) and displacing probes, we have developed a Cyclic Enzymatic Amplification Method (CEAM) for sensitive and selective detection of nucleic acids. In this design, the displacing probe is non-fluorescent on its own and cannot be digested by Exo III until displacement hybridization by a target sequence, leading to release of free non-quenched fluorophore. Because a single target sequence can lead to the release and digestion of numerous fluorophore strands from the displacing probe, a remarkable signal amplification is achieved. With this method, DNA can be detected in the picomolar range with a high selectivity and within less than 20 min.
Co-reporter:Zhi Zhu;Cuichen Wu;Haipeng Liu;Yuan Zou;Xiaoling Zhang ;Huaizhi Kang;ChaoyongJames Yang Dr.;Weihong Tan Dr.
Angewandte Chemie International Edition 2010 Volume 49( Issue 6) pp:1052-1056
Publication Date(Web):
DOI:10.1002/anie.200905570
Co-reporter:Zhi Zhu;Cuichen Wu;Haipeng Liu;Yuan Zou;Xiaoling Zhang ;Huaizhi Kang;ChaoyongJames Yang Dr.;Weihong Tan Dr.
Angewandte Chemie 2010 Volume 122( Issue 6) pp:1070-1074
Publication Date(Web):
DOI:10.1002/ange.200905570
Co-reporter:Cuichen Wu, Ling Yan, Chunming Wang, Haoxue Lin, Chi Wang, Xi Chen, Chaoyong James Yang
Biosensors and Bioelectronics 2010 Volume 25(Issue 10) pp:2232-2237
Publication Date(Web):15 June 2010
DOI:10.1016/j.bios.2010.02.030
Simple, fast and direct analysis or monitoring of significant molecules in complex biological samples is important for many biological study, clinical diagnosis and forensic investigations. Herein we highlight a general method to tailor aptamer sequence into functional subunits to design target-induced light-switching excimer sensors for rapid, sensitive and selective detection of important molecules in complex biological fluids. Our approach is to split one single strand aptamer into two pieces and each terminally labeled with a pyrene molecule while maintaining their binding affinity to target molecules. In the presence of target molecules, two aptamer fragments are induced to self-assemble to form aptamer–target complex and bring two pyrene molecules into a close proximity to form an excimer, resulting in fluorescent switching from ∼400 nm to 485 nm. With an anti-cocaine sensor, as low as 1 μM of cocaine can be detected using steady-state fluorescence assays and more importantly low picomole level of target can be directly visualized with naked eyes. Because the excimer has a long fluorescence lifetime, time-resolved measurements were used to directly detect as low as 5 μM cocaine in urine samples quantitatively without any sample pretreatment.
Co-reporter:Liang Cui, Zhi Zhu, Ninghang Lin, Huimin Zhang, Zhichao Guan and Chaoyong James Yang
Chemical Communications 2014 - vol. 50(Issue 13) pp:NaN1578-1578
Publication Date(Web):2013/12/05
DOI:10.1039/C3CC48707E
A T7 exonuclease-assisted cyclic enzymatic amplification method (CEAM) was combined with rolling circle amplification (RCA) to develop a RCA–CEAM dual amplification method for ultrasensitive detection of microRNA with excellent selectivity.
Co-reporter:Yi Xie, Xiaoyan Lin, Yishun Huang, Rujun Pan, Zhi Zhu, Leiji Zhou and Chaoyong James Yang
Chemical Communications 2015 - vol. 51(Issue 11) pp:NaN2158-2158
Publication Date(Web):2014/12/17
DOI:10.1039/C4CC08912J
Based on the protective properties of polydopamine nanospheres for DNA probes against nuclease digestion, we have developed a DNase I-assisted target recycling signal amplification method for highly sensitive and selective detection of miRNA.
Co-reporter:Huimin Zhang, Yanling Song, Yuan Zou, Yun Ge, Yuan An, Yanli Ma, Zhi Zhu and Chaoyong James Yang
Chemical Communications 2014 - vol. 50(Issue 38) pp:NaN4894-4894
Publication Date(Web):2014/03/21
DOI:10.1039/C4CC01528B
A photo-reactive functional labelling reagent, diazirine phosphoramidite, was designed and synthesized for easy and flexible site-specific labelling of oligonucleotides with the diazirine moiety. The new reagent allows facile photo-crosslinking of oligonucleotide with its interacting partner for a variety of applications, including tertiary structure determination, molecular interaction study and biomarker discovery.
Co-reporter:Xiaoyan Lin, Liang Cui, Yishun Huang, Ya Lin, Yi Xie, Zhi Zhu, Bincheng Yin, Xi Chen and Chaoyong James Yang
Chemical Communications 2014 - vol. 50(Issue 57) pp:NaN7648-7648
Publication Date(Web):2014/05/22
DOI:10.1039/C4CC02184C
Based on the protective properties of carbon nanoparticles for aptamers against the digestion of nuclease, we have developed a nuclease-assisted target recycling signal amplification method for highly sensitive detection of biomolecules, such as ATP and kanamycin. The high binding specificity between aptamers and targets leads to excellent selectivity of the assay.
Co-reporter:Xiaoyan Lin, Cheng Zhang, Yishun Huang, Zhi Zhu, Xi Chen and Chaoyong James Yang
Chemical Communications 2013 - vol. 49(Issue 65) pp:NaN7245-7245
Publication Date(Web):2013/06/18
DOI:10.1039/C3CC43224F
Based on backbone-modified molecular beacons and duplex-specific nuclease, we have developed a target recycling amplification method for highly sensitive and selective miRNA detection. The combination of a low fluorescence background of 2-OMe-RNA modified MB and nuclease-assisted signal amplification leads to ultrahigh assay sensitivity, and the powerful discriminating ability of MB enables the differentiation of highly similar miRNAs with one-base difference, both of which are of great significance to miRNA detection.
Co-reporter:Yuan Zou, Jie Chen, Zhi Zhu, Lianyu Lu, Yishun Huang, Yanling Song, Huimin Zhang, Huaizhi Kang and Chaoyong James Yang
Chemical Communications 2013 - vol. 49(Issue 77) pp:NaN8718-8718
Publication Date(Web):2013/07/29
DOI:10.1039/C3CC44188A
A pair of single-molecule photo-responsive DNA nanoscissors for DNA cleavage based on the regulation of substrate binding affinity was designed and fabricated. Compared with other DNA nanomachines, our DNA nanoscissors have the advantages of a clean switching mechanism, as well as robust and highly reversible operation.
Co-reporter:Liang Cui, Xiaoyan Lin, Ninghang Lin, Yanling Song, Zhi Zhu, Xi Chen and Chaoyong James Yang
Chemical Communications 2012 - vol. 48(Issue 2) pp:NaN196-196
Publication Date(Web):2011/10/05
DOI:10.1039/C1CC15412E
Based on graphene oxide-protected DNA probes, we have developed a cyclic enzymatic amplification method for sensitive miRNA detection in complex biological samples. By using the quenching nature of graphene oxide for multiple fluorophores, this method can distinguish highly similar miRNA sequences and detect them simultaneously.
Co-reporter:Cong Li, Ling Zhu, Zhi Zhu, Hao Fu, Gareth Jenkins, Chunming Wang, Yuan Zou, Xin Lu and Chaoyong James Yang
Chemical Communications 2012 - vol. 48(Issue 67) pp:NaN8349-8349
Publication Date(Web):2012/06/29
DOI:10.1039/C2CC32919K
In this work we studied how backbone chemical modifications, such as 2′-O-methyl, phosphorothioate, L-form nucleotides and locked nucleic acid, on G-quadruplex based DNAzymes would affect their peroxidase activity. Our results indicate that 2′-O-methyl modification facilitates the formation of a perfectly compacted parallel structure and significantly promotes peroxidase activity of G-quadruplex based DNAzymes.
Co-reporter:Chunming Wang, Zhi Zhu, Yanling Song, Hui Lin, Chaoyong James Yang and Weihong Tan
Chemical Communications 2011 - vol. 47(Issue 20) pp:NaN5710-5710
Publication Date(Web):2011/04/18
DOI:10.1039/C1CC10481K
We have constructed a novel class of light-activatable caged molecular beacons (cMBs) that are caged by locking two stems with a photo-labile biomolecular interaction or covalent bond. With the cMBs, the nucleic acid hybridization process can be easily controlled with light, which offers the possibility for a high spatiotemporal resolution study of intracellular mRNAs.
Co-reporter:Yanling Song, Tian Tian, Yuanzhi Shi, Wenli Liu, Yuan Zou, Tahereh Khajvand, Sili Wang, Zhi Zhu and Chaoyong Yang
Chemical Science (2010-Present) 2017 - vol. 8(Issue 3) pp:NaN1751-1751
Publication Date(Web):2016/12/07
DOI:10.1039/C6SC04671A
Up to 90% of cancer-related deaths are caused by metastatic cancer. Circulating tumor cells (CTCs), a type of cancer cell that spreads through the blood after detaching from a solid tumor, are essential for the establishment of distant metastasis for a given cancer. As a new type of liquid biopsy, analysis of CTCs offers the possibility to avoid invasive tissue biopsy procedures with practical implications for diagnostics. The fundamental challenges of analyzing and profiling CTCs are the extremely low abundances of CTCs in the blood and the intrinsic heterogeneity of CTCs. Various technologies have been proposed for the enrichment and single-cell analysis of CTCs. This review aims to provide in-depth insights into CTC analysis, including various techniques for isolation of CTCs with capture methods based on physical and biochemical principles, and single-cell analysis of CTCs at the genomic, proteomic and phenotypic level, as well as current developmental trends and promising research directions.
Co-reporter:Yanling Song, Weiting Zhang, Yuan An, Liang Cui, Chundong Yu, Zhi Zhu and Chaoyong James Yang
Chemical Communications 2012 - vol. 48(Issue 4) pp:NaN578-578
Publication Date(Web):2011/11/07
DOI:10.1039/C1CC15777A
We have combined an allosteric molecular beacon for target recognition and guanine-rich DNAzyme for signal amplification to develop a new platform for visual detection of nucleic acids with single-base mismatch detection capability. The fully DNA-structured platform can undergo color change in response to target DNA/RNA, which enables sensitive and selective visual detection in biological samples.
Co-reporter:Haoxue Lin, Yuan Zou, Yishun Huang, Jie Chen, Wei Yun Zhang, Zhixia Zhuang, Gareth Jenkins and Chaoyong James Yang
Chemical Communications 2011 - vol. 47(Issue 33) pp:NaN9314-9314
Publication Date(Web):2011/05/31
DOI:10.1039/C1CC12290H
We propose the use of DNAzyme as a crosslinker of hydrogel to develop a catalytic platform for the sensing of metal ions. The DNAzyme crosslinked hydrogel can undergo gel–sol transition in response to Cu2+ ions, which enables sensitive visual detection of Cu2+ by observing the release of pre-trapped AuNPs.
![Phosphoramidous acid, N,N-bis(1-methylethyl)-, 2-cyanoethyl 6-[(2-methyl-1-oxo-2-propen-1-yl)amino]hexyl ester](/data/chemimg/199400/1226983-36-3.png)
![Phosphoramidous acid, N,N-bis(1-methylethyl)-, 2-cyanoethyl 6-[(2-methyl-1-oxo-2-propen-1-yl)amino]hexyl ester](/data/chemimg/199400/1226983-36-3_b.png)
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![Phosphoramidous acid, N,N-bis(1-methylethyl)-, 2-cyanoethyl 2-[(1-oxooctadecyl)amino]-1-[[(1-oxooctadecyl)amino]methyl]ethyl ester](/data/chemimg/108500/207273-85-6_b.png)
![L-Phenylalanine,N-[[(3R)-5-chloro-3,4-dihydro-8-hydroxy-3-methyl-1-oxo-1H-2-benzopyran-7-yl]carbonyl]-](http://img.cochemist.com/ccimg/400/303-47-9.png)
![L-Phenylalanine,N-[[(3R)-5-chloro-3,4-dihydro-8-hydroxy-3-methyl-1-oxo-1H-2-benzopyran-7-yl]carbonyl]-](http://img.cochemist.com/ccimg/400/303-47-9_b.png)
![Methyl (3s,4r)-3-benzoyloxy-8-methyl-8-azabicyclo[3.2.1]octane-4-carboxylate](http://img.cochemist.com/ccimg/100/50-36-2.png)
![Methyl (3s,4r)-3-benzoyloxy-8-methyl-8-azabicyclo[3.2.1]octane-4-carboxylate](http://img.cochemist.com/ccimg/100/50-36-2_b.png)