Jiangsu Institute of Cancer Prevention and Cure

Nanjing, China

Jiangsu Institute of Cancer Prevention and Cure

Nanjing, China
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Lin D.,Nanjing University | Wu J.,Nanjing University | Ju H.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure
Biosensors and Bioelectronics | Year: 2014

Nanogold functionalized mesoporous carbon foam (Au/MCF) coupling with a signal amplification by C-Au synergistic silver enhancement was designed for sensitive electrochemical immunosensing of biomarker. The Au/MCF was prepared by in situ growth of nanogold on carboxylated MCF and used as a tracing tag to label signal antibody via the inherent interaction between protein and nanogold. The immunosensor was prepared by covalently immobilizing capture antibody on an electrochemically reduced graphene oxide/chitosan film modified glassy carbon electrode. Through a sandwich-type immunoreaction, Au/MCF tags were captured on the immunoconjugates to induce a silver deposition process. The electrochemical stripping signal of the deposited silver was used to monitor the immunoreaction. The Au/MCF-mediated silver enhancement along with the graphene-promoted electron transfer led to high detection sensitivity of carcinoembryonic antigen. Under optimal conditions, the proposed immunoassay method showed wide linear range from 0.05pgmL-1 to 1ngmL-1 and a detection limit down to 0.024pgmL-1. The newly designed amplification strategy holds great potential for ultrasensitive electrochemical biosensing of other analytes. © 2013 Elsevier B.V.


Dong H.,Nanjing University | Ding L.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure | Ji H.,Nanjing University | Ju H.,Nanjing University
Biomaterials | Year: 2011

A simple nanocarrier of polyethylenimine-grafted graphene nanoribbon (PEI-g-GNR) was proposed as an effective gene vector. The GNR was formed by longitudinally unzipping multiwalled carbon nanotubes (MWCNTs), and treated with strong acids and sonication to obtain surface carboxylic acid groups for graft of PEI via electrostatic assembly. The PEI-g-GNR appeared to protect locked nucleic acid modified molecular beacon (LNA-m-MB) probes from nuclease digestion or single-strand binding protein interaction, thus could be used as a nanocarrier of the probes for more efficient transfection of cells than PEI or PEI-g-MWCNTs due to the large surface area of the GNR and high charge density of PEI. The cytotoxicity and apoptosis induced by the PEI-g-GNR were negligible under optimal transfection conditions. Combining with the remarkable affinity and specificity of LNA to microRNA (miRNA), a delivery system by the LNA-m-MB/PEI-g-GNR was proposed for effectively transferring LNA-m-MB into the cells to recognize the target miRNA. Using HeLa cells as model, a method for detection of miRNA in single cell was developed. These results suggested that PEI-g-GNR would be a promising nonviral vector for in situ detection of gene in cytoplasm and gene therapy in clinical application. © 2011 Elsevier Ltd.


Ji H.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure | Lei J.,Nanjing University | Ju H.,Nanjing University
Analytical Chemistry | Year: 2012

An ultrasensitive protocol for electrochemical detection of DNA is designed with quantum dots (QDs) as a signal tag by combining the template enhanced hybridization process (TEHP) and rolling circle amplification (RCA). Upon the recognition of the molecular beacon (MB) to target DNA, the MB hybridizes with assistants and target DNA to form a ternary "Y-junction". The target DNA can be dissociated from the structure under the reaction of nicking endonuclease to initiate the next hybridization process. The template enhanced MB fragments further act as the primers of the RCA reaction to produce thousands of repeated oligonucleotide sequences, which can bind with oligonucleotide functionalized QDs. The attached signal tags can be easily read out by square-wave voltammetry after dissolving with acid. Because of the cascade signal amplification and the specific TEHP and RCA reaction, this newly designed protocol provides an ultrasensitive electrochemical detection of DNA down to the attomolar level (11 aM) with a linear range of 6 orders of magnitude (from 1 × 10-17 to 1× 10-11 M) and can discriminate mismatched DNA from perfect matched target DNA with high selectivity. The high sensitivity and specificity make this method a great potential for early diagnosis in gene-related diseases. © 2012 American Chemical Society.


Lin D.,Nanjing University | Wu J.,Nanjing University | Wang M.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure | Ju H.,Nanjing University
Analytical Chemistry | Year: 2012

A triple signal amplification strategy was designed for ultrasensitive immunosensing of cancer biomarker. This strategy was achieved using graphene to modify immunosensor surface for accelerating electron transfer, poly(styrene-co-acrylic acid) microbead (PSA) carried gold nanoparticles (AuNPs) as tracing tag to label signal antibody (Ab 2) and AuNPs induced silver deposition for anodic stripping analysis. The immunosensor was constructed by covalently immobilizing capture antibody on chitosan/ electrochemically reduced graphene oxide film modified glass carbon electrode. The in situ synthesis of AuNPs led to the loading of numerous AuNPs on PSA surface and convenient labeling of the tag to Ab 2. With a sandwich-type immunoreaction, the AuNPs/PSA labeled Ab 2 was captured on the surface of an immunosensor to further induce a silver deposition process. The electrochemical stripping signal of the deposited silver nanoparticles in KCl was used to monitor the immunoreaction. The triple signal amplification greatly enhanced the sensitivity for biomarker detection. The proposed method could detect carcinoembryonic antigen with a linear range of 0.5 pg mL -1 to 0.5 ng mL -1 and a detection limit down to 0.12 pg mL -1. The immunosensor exhibited good stability and acceptable reproducibility and accuracy, indicating potential applications in clinical diagnostics. © 2012 American Chemical Society.


Xu Q.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure | Lei J.,Nanjing University | Leng C.,Nanjing University | Ju H.,Nanjing University
Chemistry - A European Journal | Year: 2012

This work designed a simple, sensitive, and low-cost immunosensor for the detection of protein marker by using a carbon sphere/gold nanoparticle (CNS/AuNP) composite as an electrochemical label. The nanoscale carbon spheres, prepared with a hydrothermal method by using glucose as raw material, were used to load AuNPs for labeling antibody by electrostatic interaction, which provided a feasible pathway for electron transfer due to the remarkable conductivity. The disposable immunosensor was constructed by coating a polyethylene glycol (PEG) film on a screen-printed carbon-working electrode and then immobilizing capture antibody on the film. With a sandwich-type immunoassay format, the analyte and then the CNS/AuNP-labeled antibody were successively bound to the immunosensor. The bound AuNPs were finally electro-oxidized in 0.1M HCl to produce AuCl 4 - for differential pulse voltammetric (DPV) detection. The high-loading capability of AuNPs on CNS for the sandwich-type immunorecognition led to obvious signal amplification. By using human immunoglobulin G (IgG) as model target, the DPV signal of AuNPs after electro-oxidized at optimal potential of +1.40 V for 40 s showed a wide linear dependence on the logarithm of target concentration ranging from 10 pgmL -1 to 10 ngmL -1. The detection limit was around 9 pgmL -1. The immunosensor showed excellent analytical performance with cost effectivity, good fabrication reproducibility, and acceptable precision and accuracy, providing significant potential application in clinical analysis. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Zong C.,Nanjing University | Wu J.,Nanjing University | Wang C.,Nanjing University | Ju H.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure
Analytical Chemistry | Year: 2012

A sensitive chemiluminescence (CL) imaging immunoassay method for detection of multiple tumor markers with high throughput, easy operation, and low cost was developed. The immunosensor array was prepared by covalently immobilizing capture antibodies on corresponding sensing sites on a silanized disposable glass chip. Gold nanoparticle-based bioconjugates with a high molar ratio of horseradish peroxidase (HRP) to detection antibodies were used for signal amplification. Under a sandwich immunoassay, the CL signals triggered by HRP captured on each sensing cell were collected by a charge-coupled device for simultaneous measurement of biomarkers and combination diagnosis of certain tumors. As a proof of concept, the immunosensor array was applied to detect α-fetoprotein, carcinoma antigen 125, carbohydrate antigen 153, and carcinoembryonic antigen and to screen patients with liver, breast, or ovarian cancers. This method showed wide linear ranges over 5 orders of magnitude and much lower detection limits than previously reported multiplexed immunoassays. The high throughput and acceptable stability, reproducibility, and accuracy showed good applicability of the proposed multiplex CL imaging immunoassay in clinical diagnosis. © 2012 American Chemical Society.


Lai G.,Nanjing University | Wu J.,Nanjing University | Ju H.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure
Advanced Functional Materials | Year: 2011

A streptavidin-functionalized silver-nanoparticle-enriched carbon nanotube (CNT/Ag NP) is designed as trace tag for ultrasensitive multiplexed measurements of tumor markers using a disposable immunosensor array. The CNT/Ag NP nanohybrid is prepared by one-pot in situ deposition of Ag NPs on carboxylated CNTs. The nanohybrid is functionalized with streptavidin via the inherent interaction between the protein and Ag NPs for further linkage of biotinylated signal antibodies to obtain tagged antibodies. The functionalization process greatly improves the dispersibility of the nanohybrid in water. The immunosensor array is prepared by covalently immobilizing capture antibodies on chitosan-modified screen-printed carbon electrodes. Through a sandwich-type immunoreaction on the immunosensor array, numerous Ag NPs are captured onto every single immunocomplex and are further amplified by a subsequent Ag NP-promoted deposition of silver from a silver enhancer solution to obtain the sensitive electrochemical-stripping signal of the Ag NPs. Using carcinoembryonic antigen and α-fetoprotein as model analytes, this proposed multiplexed immunoassay method shows acceptable precision and wide linear ranges over four orders of magnitude with detection limits down to 0.093 and 0.061 pg mL -1, respectively. The assay results of serum samples with the proposed method are in acceptable agreement with the reference values. The newly designed strategy and the functionalized tag avoid cross-talk and the requirement of deoxygenation for electrochemical immunoassay, and thus provide a promising potential in clinical application. A streptavidin-functionalized silver-nano-particle-enriched carbon nanotube is designed as a trace tag for ultrasensitive multiplexed detection of tumor markers by combination with silver-deposition enhancement and electrochemical stripping analysis of silver nanoparticles on a disposable immunosensor array. The excellent analytical performance of this method demonstrates that the functionalized nanohybrid is a promising and versatile amplified probe for application in biotechnology. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Lai G.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure | Wu J.,Nanjing University | Leng C.,Nanjing University | Ju H.,Nanjing University
Analytical Chemistry | Year: 2011

A novel ultrasensitive multiplexed immunoassay method was developed by combining alkaline phosphatase (ALP)-labeled antibody functionalized gold nanoparticles (ALP-Ab/Au NPs) and enzyme-Au NP catalyzed deposition of silver nanoparticles at a disposable immunosensor array. The immunosensor array was prepared by covalently immobilizing capture antibodies on chitosan modified screen-printed carbon electrodes. After sandwich-type immunoreactions, the ALP-Ab/Au NPs were captured on an immunosensor surface to catalyze the hydrolysis of 3-indoxyl phosphate, which produced an indoxyl intermediate to reduce Ag+. The silver deposition process was catalyzed by both ALP and Au NPs, which amplified the detection signal. The deposited silver was then measured by anodic stripping analysis in KCl solution. Using human and mouse IgG as model analytes, this multiplexed immunoassay method showed wide linear ranges over 4 orders of magnitude with the detection limits down to 4.8 and 6.1 pg/mL, respectively. Acceptable assay results for practical samples could be obtained. The newly designed strategy avoided cross talk and the need of deoxygenation for the electrochemical immunoassay and, thus, provided a promising potential in clinical applications. © 2011 American Chemical Society.


Xu J.,Nanjing University | Wu J.,Nanjing University | Zong C.,Nanjing University | Ju H.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure
Analytical Chemistry | Year: 2013

Manganese porphyrin (MnTMPyP)-dsDNA complex was reported as an excellent mimicking enzyme of peroxidase. It possessed high catalytic activity and much quicker catalytic kinetics and better stability with exposure to light irradiation and high temperature than both horseradish peroxidase and hemin/G-quadruplex DNAzyme. The groove binding of MnTMPyP to the dsDNA scaffold efficiently maintained the catalytic activity of the MnTMPyP center and improved its stability. By combining with an isothermal hybridization chain reaction (HCR) and in situ formation of MnTMPyP-dsDNA, a highly efficient chemiluminescent (CL) immunosensing method was proposed. After a sandwich immunoreaction, a biotinylated DNA strand, which was bound to biotinylated signal antibody by streptavidin, triggered the HCR and growth of MnTMPyP-dsDNA on the immunocomplex. The in situ, HCR-assisted enzyme formation brought numerous enzymatic catalytic centers, MnTMPyP, on the immunocomplex, resulting in significant CL signal amplification and highly sensitive CL detection. Using carcinoembryonic antigen as the model target, the proposed CL immunoassay method showed a wide linear range from 10 pg/mL to 100 ng/mL with a detection limit of 6.8 pg/mL. The new MnTMPyP-dsDNA complex could be conveniently synthesized, functionalized, and combined with DNA amplification strategies, showing a promising potential in bioanalysis and other relative fields. © 2013 American Chemical Society.


Dong H.,Nanjing University | Zhu Z.,Nanjing University | Ju H.,Nanjing University | Yan F.,Jiangsu Institute of Cancer Prevention and Cure
Biosensors and Bioelectronics | Year: 2012

An ultrasensitive electrochemical DNA biosensor was constructed by assembling probe labeled gold nanoparticles (ssDNA-AuNP) on electrochemically reduced graphene oxide (ERGO) modified electrode with thiol group tagged (GT) DNA strand (d(GT) 29SH) and coupling with horseradish peroxidase (HRP) functionalized carbon sphere (CNS) as tracer. The heteronanostructure formed on the biosensor surface appeared relatively good conductor for accelerating the electron transfer, while the HRP tagged CNS provided dual signal amplification for electrochemical biosensing. The triplex signal amplification strategy produced an ultrasensitive electrochemical detection of DNA down to attomolar level (5aM) with a linear range of 5 orders of magnitude (from 1×10 -17M to 1×10 -13M), and appeared high selectivity to differentiate single-base mismatched and three-base mismatched sequences of DNA. The proposed approach provided a simple and reliable method for DNA detection with high sensitivity and specificity, indicating promising application in bioanalysis and biomedicine. © 2012 Elsevier B.V.

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