Guonan Chen

Gold nanoparticles-decorated amine-terminated poly(amidoamine) dendrimer for sensitive electrochemical immunoassay of brevetoxins in food samples

A sensitive electrochemical immunosensor for the fast screening of brevetoxin B (BTX-2) in food samples was developed by means of immobilizing BTX-2-bovine serum albumin conjugate (BTX-2-BSA) on the gold nanoparticles-decorated amine-terminated poly(amidoamine) dendrimers (AuNP-PAADs). The presence of gold nanoparticles greatly improved the conductivity of the PAADs, and three-dimensional PAADs increased the surface coverage of the biomolecules on the electrode. Under optimal conditions, three types of immunosensor, i.e. with AuNPs, PAADs, or AuNP-PAADs, were used for the determination of BTX-2 in a competitive-type immunoassay format using horseradish peroxidase-labeled anti-BTX antibodies (HRP-anti-BTX-2) as trace in the H(2)O(2)-o-phenylenediamine (o-PD) system. A low detection limit (LOD) of 0.01 ng/mL and a wide dynamic working linear range of 0.03-8 ng/mL BTX-2 using AuNP-PAADs as matrices were obtained in comparison with those of only using AuNP or PAADs. Intra-batch assay precision was substantially improved by resorting to the AuNP-PAADs manifold. The proposed method features unbiased identification of negative (blank) and positive samples. No significant differences were encountered in the analysis of the spiking real samples between the electrochemical immunosensor and liquid chromatography for the determination of BTX-2. Importantly, this method provided a biocompatible immobilization and a promising immunosensing platform for analytes with small molecules in the analysis and detection of food safety.

Biocompatible electrochemiluminescent biosensor for choline based on enzyme/titanate nanotubes/chitosan composite modified electrode.

A new biocompatible ECL biosensor based on enzyme/titanate nanotubes/chitosan composite film was developed for the determination of analytes in biological samples. In the fabrication of the new ECL biosensor, biocompatible titanate nanotubes (TNTs) and a model enzyme, i.e., choline oxidase (ChOX), were immobilized on a chitosan modified glassy carbon electrode (GCE) via electrostatic adsorption and covalent interaction, respectively. By this ECL biosensor, choline was enzymatically oxidized to hydrogen peroxide and detected by a sensitive luminol ECL system. The use of TNTs not only provided a biocompatible microenvironment for the immobilized enzyme, which resulted in an excellent stability and long lifetime of the ECL biosensor, but also exhibited great enhancement towards luminol ECL and thus led to a significant improvement in sensitivity of ECL biosensor. Satisfactory results were obtained when employing this biosensor in assaying the total choline in milk samples. The work would provide a common platform to develop various sensitive, selective and biocompatible ECL biosensors based on using enzyme/TNTs/CHIT composite films.

GC/MS-based metabolomics reveals fatty acid biosynthesis and cholesterol metabolism in cell lines infected with influenza A virus

Metabolomics is the downstream of systems biology and has drawn significant interest for studying the metabolic networks from cells to organisms. To profile the metabolites in two different cell lines (A549 and AGS) infected with influenza A virus, gas chromatography coupled with mass spectrometry (GC/MS) was employed. Some differentiating metabolites in the cell lines were tentatively identified using reference library, interpreted and visualized by applying principal components analysis (PCA) and cluster heat map. Consequently, metabolic flux profiling allowed the differentiation of fatty acid biosynthesis and cholesterol metabolism during viral replication in the cell lines. The change in fatty acid turnover was also observed. Metabolomics investigation also revealed the different responses between A549 and AGS cell lines to the virus infection. From the pattern recognition results, AGS cell line might be more susceptible to influenza A virus. Regarding the fact that AGS is a poorly differentiated gastric adenocarcinoma cell line whereas A549 is a relatively differentiated lung tumor one, it is speculated that viral replication might be associated with the cell differentiations.

Flow injection chemiluminescent detection of trace Co(II) with dibromoalizarin violet-H2O2-CTMAB system

Chemiluminescence was observed when dibromoalizarin violet was oxidized with hydrogen peroxide in alkaline solution. Trace amounts of Co (II) catalysed this CL reaction strongly, especially in the presence of the cationic surfactant cetyltrimethylammonium bromide, and the CL intensity was proportional to the concentration of Co(II). A flow injection system with CL detection was established to investigate this CL system. The optimum conditions for this CL reaction were investigated in detail, and the optimized flow injection parameters were determined by the modified simplex method. A CL analytical method for determination of ultratrace amounts of Co (II) was developed with a detection limit of 4 pg/mL. It was used for analysis of natural water samples, and the results compare very well with those from GFAAS. A possible mechanism for this CL reaction is proposed on the basis of studying CL spectra, absorption spectra, fluorescent spectra and HMO treatment for the reagent molecule. The effects of various types of surfactants on CL reaction are also discussed.

Electrochemical behavior of thionine at titanate nanotubes-based modified electrode: A sensing platform for the detection of trichloroacetic acid

Titanate nanotubes (TNTs) have some unique and seductive properties, such as good biocompatibility, negative charge after treating with weak bases and supply of special reaction vessel for analyte due to their hydroxyls and multilayered structures. Herein, titanate nanotubes were firstly self-assembled on the chitosan (CHIT) membrane modified electrode. Then thionine, as a model electrochemical probe, was firmly immobilized onto the titanate nanotubes (TNTs)-based sensing interface. This strategy provided a new route to immobilize the redox mediator onto the sensing interface. Based on which, trichloroacetic acid (TCA) was found to be able to accelerate the electron transmission rate and improve the electrochemical behavior of thionine. Therefore, a wide linear range for the detection of TCA was estimated to be from 6microM to 1.5mM with high sensitivity and good selectivity.

Micro-plate Chemiluminescence Enzyme Immunoassay for Clinical Determination of Progesterone in Human Serum

Abstract A high-throughput, simple and rapid chemiluminescence enzyme immunoassay (CLEIA) was developed for the clinical determination of progesterone in human serum, using luminol-hydrogen peroxide as chemiluminescence system catalyzed by horseradish peroxidase (HRP). The solid phase of anti-progesterone antibody was prepared by immunoreaction between anti-progesterone polyclonal antibody and donkey anti-rabbit IgG, i.e. second antibody, which had been physically absorbed on the wells of polystyrene microplate and was used as a universal solid phase. The effect of various factors, such as the dilution of immunoreagents, chemiluminescence substrate, chemiluminescence reaction time and incubation condition were examined and optimized. The optimal dilutions of anti-progesterone antibody and HRP-progesterone conjugate were 1:10000 and 1:15000, respectively. The II substrate was chosen and the luminescence was determined after 10 min incubation. The immunoreacted sample was incubated in water bath at 37°C for 1 h. The assay was evaluated with sensitivity as low as 0.08 μg l −1 . The relative standard deviation (RSD) was less than 15% for both intra- and inter-assay precision. The recoveries of three different spiked concentration samples were 101%, 101% and 94.4%, respectively. This proposed method has been successfully applied to the clinical evaluation of progesterone in 36 human sera. The results showed a good correlation with the accredited radioimmunoassay (RIA) with a correlative coefficient of 0.9502.

Determination of Free Triiodothyronine by Chemiluminescent Enzyme Immunoassay

Abstract The methodology based on chemiluminescent enzyme immunoassay was established for the determination of free Triiodothyronine (FT3). The HRP-luminol-H2O2 chemiluminescent system with high sensitivity was chosen as the detection system. The linear range was 0.90–80 pg ml−1. Under the selected conditions, the CVs of intra- and inter- assays were less than 15%. When the method was applied to detect free triiodothyronine in human serum, the diagnostic accordance rate of the method for hyperthyroidism was 83.3%. Compared with the commercial kit, the correlation coefficient was 0.9005. All the conditions chosen were to maintain the equilibrium between the free and the bound hormones to improve the validity of the experimental result.

Optimization of Gradient Elution in UPLC: A Core Study on the Separation of Homoserine Lactones Produced by Bukholderia Ubonensis and Structure Confirmation with Ultra High Resolution Mass Spectrometry

Abstract Speed of analysis is a key factor in liquid chromatography. One possible way is to decrease the particle size and thus increase the pressure (ultra high pressure liquid chromatography, UHPLC). For the separation of a complex mixture, the application of gradient elution techniques is expedient. From a practical point of view, it has to be questioned how the high pressure influences the separation parameters at gradient mode. In our study we investigated the following chromatographic parameters: peak width versus temperature, peak width versus gradient time, retention time versus temperature, and retention factor versus flow rate. It was shown that even in sample volume overload conditions the gradient system worked in a reproducible way; the maximum possible sample volume (20 µL) was injected to get the lowest possible limit of detection. Increasing the flow rate, the peak width increased only moderately, and the same was found when the temperature was decreased. However, at 60°C the signal to noise ratio decreased in a small compass showing the thermal disequilibrium between the column outlet and the detector cell, which might have caused extra noise. Finally, a practical separation was presented in which the UHPLC can be applied for the determination of N‐acylhomoserine lantones (AHLs), which have a relatively low UV absorption at low wavelength. The detection limit was low enough for practical applications because of the low dispersion due to a small diameter stationary phase and the low dead volume in the column. The UPLC method was applied as a first screening for AHLs and the results were confirmed by fourier transformation ion cyclotron resonance mass spectrometry (FTICR‐MS), which has a high selectivity.