Nengqin Jia

Chitosan-coated magnetic nanoparticles as carriers of 5-Fluorouracil: Preparation, characterization and cytotoxicity studies

The chitosan-coated magnetic nanoparticles (CS MNPs) were prepared as carriers of 5-Fluorouracil (CS-5-Fu MNPs) through a reverse microemulsion method. The characteristics of CS-5-Fu MNPs were determined by using transmission electron microscopy (TEM), FTIR spectroscopy and vibrating-sampling magnetometry (VSM). It was found that the synthesized CS-5-Fu MNPs were spherical in shape with an average size of 100+/-20 nm, low aggregation and good magnetic responsivity. Meanwhile, the drug content and encapsulation rate of the nanoparticles was 16-23% and 60-92%, respectively. These CS-5-Fu MNPs also demonstrated sustained release of 5-Fu at 37 degrees C in different buffer solutions. The cytotoxicity of CS-5-Fu MNPs towards K562 cancer cells was investigated. The result showed that CS-5-Fu MNPs retained significant antitumor activities. Additionally, it was observed that the FITC-labeled CS-5-Fu MNPs could effectively enter into the SPCA-1 cancer cells and induced cell apoptosis.

Improved EIS performance of an electrochemical cytosensor using three-dimensional architecture Au@BSA as sensing layer.

An ultrasensitive electrochemical cytosensor for quantitative determination of carcinoembryonic antigen (CEA)-positive tumor cells was developed using three-dimensional (3D) architecture Au@BSA microspheres as sensing layer with the conjugation of targeting molecule monoclonal anti-CEA antibody (anti-CEA). The prepared Au@BSA microspheres exhibited satisfactory biocompatibility for cell proliferation via evaluation from thiazolyl blue tetrazolium bromide (MTT) assay, providing a suitable platform for cell adhesion study. Attributed to the excellent electroconductivity of Au@BSA, amplified electrochemical signals could be obtained and resulted in the greatly enhanced detection sensitivity. Electrochemical testing techniques including electrochemical impedance spectroscopy (EIS), differential pulse voltammetry (DPV), and cyclic voltammetry (CV) were applied to assess the optimal conditions, specificity, and detection performance of as-fabricated cytosensor. The attachment of CEA-positive BXPC-3 cells onto the anti-CEA immobilized sensing layer led to the increased EIS responses, which changed linearly in the cell concentration range from 5.2 × 10(1) to 5.2 × 10(7) cells mL(-1) with a detection limit of 18 cells mL(-1). This proposed cytosensing strategy revealed high specificity to CEA-positive cells, acceptable intra-assay precision, excellent fabrication reproducibility with the RSD of 3.5%, and good stability owing to the outside BSA biocompatible layer, developing a promising technique for early monitoring of tumor cells at a lower level.

Selective and Ratiometric Fluorescent Trapping and Quantification of Protein Vicinal Dithiols and in Situ Dynamic Tracing in Living Cells

Protein vicinal dithiols play fundamental roles in intracellular redox homeostasis due to their involvement in protein synthesis and function through the reversible vicinal dithiol oxidation to disulfide. To provide quantitative information about the global distribution and dynamic changes of protein vicinal dithiols in living cells, we have designed and synthesized a ratiometric fluorescent probe (VTAF) for trapping of vicinal dithiol-containing proteins (VDPs) in living cells. VTAF exhibits a ratiometric fluorescence signal upon single excitation, which enables self-calibration of the fluorescence signal and quantification of endogenous vicinal dithiols of VDPs. Its potential for in situ dynamic tracing of changes of protein vicinal dithiols under different cellular redox conditions was exemplified. VTAF facilitated the direct observation of subcellular distribution of endogenous VDPs via ratiometric fluorescence imaging and colocalization assay. And the results suggested that there are abundant VDPs in mitochondria. Moreover, some redox-sensitive VDPs are also present on cell surface which can respond to redox stimulus. This ratiometric fluorescence technique presents an important extension to previous fluorescence intensity-based probes for trapping and quantifying protein vicinal dithiols in living cells, as well as its visible dynamic tracing of VDPs.

Ag@BSA Core/Shell Microspheres As an Electrochemical Interface for Sensitive Detection of Urinary Retinal-Binding Protein

The level of urinary retinol-binding protein (RBP) can be estimated as a significant index of renal tubular injury. In this work, we used Ag@BSA microspheres as a sensing interface to cross-link RBP monoclonal antibody (RBP mAb) via glutaraldehyde for sensitive detection of RBP. The Ag@BSA microspheres covered on a Au electrode could provide a larger surface area and multifunctional substrate for the effective immobilization of RBP mAb, and the outside BSA layer acted as a biocompatible support to maintain the bioactivity and stability of immobilized immunogen. Electrochemical measurements containing electrochemical impedance spectroscopy (EIS) and differential pulse voltammetry (DPV) were employed to evaluate the analytical performance of the fabricated immunosensor and a higher detection sensitivity was obtained by DPV attributed to the excellent electrical conductivity of Ag@BSA which could enhance the peak current response. This immunosensor had a best detection limit (DL) of 18 ng mL(-1) and a linear response range between 50 and 4500 ng mL(-1). The proposed approach showed high specificity for RBP detection, acceptable reproducibility with an RSD of 5.6%, and good precision with the RSD of 4.5% and 6.3% at the RBP concentrations of 500 and 1500 ng mL(-1). Compared with the ELISA method by analyzing real urine samples from a patient, this immunosensor revealed acceptable accuracy with a relative deviation lower than 6.5%, indicating a potential alternative method for RBP detection in clinical diagnosis.

Functionalized halloysite nanotube-based carrier for intracellular delivery of antisense oligonucleotides

Halloysites are cheap, abundantly available, and natural with high mechanical strength and biocompatibility. In this paper, a novel halloysite nanotube [HNT]-based gene delivery system was explored for loading and intracellular delivery of antisense oligodeoxynucleotides [ASODNs], in which functionalized HNTs [f-HNTs] were used as carriers and ASODNs as a therapeutic gene for targeting survivin. HNTs were firstly surface-modified with γ-aminopropyltriethoxysilane in order to facilitate further biofunctionalization. The f-HNTs and the assembled f-HNT-ASODN complexes were characterized by transmission electron microscopy [TEM], dynamic light scattering, UV-visible spectroscopy, and fluorescence spectrophotometry. The intracellular uptake and delivery efficiency of the complexes were effectively investigated by TEM, confocal microscopy, and flow cytometry. In vitro cytotoxicity studies of the complexes using MTT assay exhibited a significant enhancement in the cytotoxic capability. The results exhibited that f-HNT complexes could efficiently improve intracellular delivery and enhance antitumor activity of ASODNs by the nanotube carrier and could be used as novel promising vectors for gene therapy applications, which is attributed to their advantages over structures and features including a unique tubular structure, large aspect ratio, natural availability, rich functionality, good biocompatibility, and high mechanical strength.

A novel solid-state electrochemiluminescence sensor for melamine with Ru(bpy)32+/mesoporous silica nanospheres/Nafion composite modified electrode

A novel melamine electrochemiluminescence (ECL) sensor was developed based on mesoporous SiO(2) nanospheres/Ru(bpy)(3)(2+)/Nafion modified electrodes. The homogeneous mesoporous silica nanospheres, synthesized using modified Stöber sol-gel process, were characterized by Field Emission Scanning Electron Microscopy (FE-SEM), Transmission Electron Microscopy (TEM) and Brunauer-Emmett-Teller (BET). The ECL and electrochemistry of the modified electrodes were investigated with tri-n-propylamine (TPA) as the coreactant. Furthermore, the mesporous SiO(2) nanospheres/Ru(bpy)(3)(2+)-based modified electrodes were used for ECL determination of melamine. The analytical performances of this ECL sensor for melamine based on its enhancement ECL emission of Ru(bpy)(3)(2+) were investigated. The results indicated that the sensor exhibited excellent performance during melamine determination with a wide linear range (7.81×10(-9)-5×10(-6) M), low detection limit (2.6×10(-9) M). The high sensitivity and stability mainly resulted from the high surface area and special structure of the mesoporous silica nanospheres. The proposed ECL approach was used to analyze the melamine content in powdered milk with satisfactory results.

Bio-mimetically synthesized Ag@BSA microspheres as a novel electrochemical biosensing interface for sensitive detection of tumor cells

The use of a novel cytosensor, comprised of bio-mimetically synthesized Ag@BSA composite microspheres, for the detection of KB cells (a model system) is described. The Ag@BSA composite microspheres were immobilized on Au electrodes via Au-thiol bonds. Scanning electron microscopy (SEM), atomic force microscopy (AFM) and transmission electron microscopy (TEM) images revealed that the Ag@BSA were well-dispersed microspheres with an average diameter of 500 nm, including the monolayer of BSA. The immobilization of Ag@BSA composite microspheres onto Au electrodes is thought to increase the electrode surface area and accelerate the electron transfer rate while providing a highly stable matrix for the convenient conjugation of target molecules (such as folic acid) and the prolonged incubation of cells. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) studies showed that the fabricated cytosensor was able to detect KB cells ranging from 6.0×10(1) to 1.2×10(8) cells mL(-1) with a lower detection limit of 20 cells mL(-1). Due to its facile synthesis, high stability and reproducibility and cytocompatibility, the novel cytosensor described here could find multifarious uses in applications, such as cancer diagnosis, drug screening and cell adhesion studies.

Highly sensitive electrochemical impedance spectroscopy immunosensor for the detection of AFB1 in olive oil

Aflatoxin produced by Aspergillus flavus and Aspergillus parasiticus are commonly found in olive and its derivatives. Aflatoxin B1 (AFB1) is a predominant toxin detected abundantly and has been implicated in the etiology of human hepatocellular carcinoma. This study proposes a sensitive and convenient electrochemical impedance spectroscopy (EIS) method for determining AFB1 by MWCNTs/RTIL composite films-based immunosensor. The calibration curve for AFB1 was linear in the range of 0.1-10ngmL(-1) with the limit of detection (LOD) 0.03ngmL(-1). The presence of MWCNTs warrant fast electron transfer, and the ionic liquid provides a benign microenvironment for antibody. The experimental parameters, such as pH and incubating time, have been investigated and optimized. Furthermore, the detection of AFB1 is presented to test this method after extracted from olive oils. It can be anticipated that this method would be used for the detection of AFB1 in various agriculture products and vegetable oils.

Sensitive ECL immunosensor for detection of retinol-binding protein based on double-assisted signal amplification strategy of multiwalled carbon nanotubes and Ru(bpy)32+ doped mesoporous silica nanospheres

A novel electrochemiluminescence (ECL) strategy based on the sandwich-type immunosensor for sensitive detection of retinol-binding protein (RBP) was developed. The primary antibody anti-RBP was immobilized onto multiwalled carbon nanotubes (MWCNTs), which have large surface area and high electrical conductivity. The RBP antigen and Ru-Nafion@SiO2-labeled secondary antibody were then successively conjugated to form sandwich-type immunocomplexes through the specific interaction between antigen and antibody. The ECL signal amplification was significantly improved due to the synergistic effect of MWCNTs and mesoporous silica nanospheres (mSiO2). The developed ECL immunosensor exhibited high sensitivity and specificity for the detection of RBP and responded linearly to the clinically-relevant concentration of RBP from 78 to 5000 ng mL(-1). Moreover, the MWCNT-based ECL immunosensor displayed excellent stability and reproducibility, as well as successfully achieved the detection of RBP in patient urine samples with desirable results. The present work provided a promising technique for the clinical screening of RBP and point-of-care diagnostics.

Magnetic Fe3O4-Reduced Graphene Oxide Nanocomposites-Based Electrochemical Biosensing

An electrochemical biosensing platform was developed based on glucose oxidase (GOx)/Fe3O4- reduced graphene oxide (Fe3O4-RGO) nanosheets loaded on the magnetic glassy carbon electrode (MGCE). With the advantages of the magnetism, conductivity and biocompatibility of the Fe3O4-RGO nanosheets, the nanocomposites could be facilely adhered to the electrode surface by magnetically controllable assembling and beneficial to achieve the direct redox reactions and electrocatalytic behaviors of GOx immobilized into the nanocomposites. The biosensor exhibited good electrocatalytic activity, high sensitivity and stability. The current response is linear over glucose concentration ranging from 0.05 to 1.5 mM with a low detection limit of 0.15 μM. Meanwhile, validation of the applicability of the biosensor was carried out by determining glucose in serum samples. The proposed protocol is simple, inexpensive and convenient, which shows great potential in biosensing application.