Fanggui Ye

Determination of uric acid in human urine and serum by capillary electrophoresis with chemiluminescence detection

A simple and sensitive method based on capillary electrophoresis (CE) with chemiluminescence (CL) detection has been developed for the determination of uric acid (UA). The sensitive detection was based on the enhancement effect of UA on the CL reaction between luminol and potassium ferricyanide (K3[Fe(CN)6]) in alkaline solution. A laboratory-built reaction flow cell and a photon counter were deployed for the CL detection. Experimental conditions for CL detection were studied in detail to achieve a maximum assay sensitivity. Optimal conditions were found to be 1.0 x 10(-4) M luminol added to the CE running buffer and 1.0 x 10(-4) M K3[Fe(CN)6] in 0.2 M NaOH solution introduced postcolumn. The proposed CE-CL assay showed good repeatability (relative standard deviation [RSD]=3.5%, n=11) and a detection limit of 3.5 x 10(-7) M UA (signal/noise ratio [S/N]=3). A linear calibration curve ranging from 6.0 x 10(-7) to 3.0 x 10(-5) M UA was obtained. The method was evaluated by quantifying UA in human urine and serum samples with satisfactory assay results.

Magnetic Bead-Sensing-Platform-Based Chemiluminescence Resonance Energy Transfer and Its Immunoassay Application

A competitive immunoassay based on chemiluminescence resonance energy transfer (CRET) on the magnetic beads (MBs) is developed for the detection of human immunoglobulin G (IgG). In this protocol, carboxyl-modified MBs were conjugated with horseradish peroxidase (HRP)-labeled goat antihuman IgG (HRP-anti-IgG) and incubated with a limited amount of fluorescein isothiocyanate (FITC)-labeled human IgG to immobilize the antibody-antigen immune complex on the surface of the MBs, which was further incubated with the target analyte (human IgG) for competitive immunoreaction and separated magnetically to remove the supernatant. The chemiluminescence (CL) buffer (containing luminol and H(2)O(2)) was then added, and the CRET from donor luminol to acceptor FITC in the immunocomplex on the surface of MBs occured immediately. The present protocol was evaluated for the competitive immunoassay of human IgG, and a linear relationship between CL intensity ratio (R = I(425)/I(525)) and human IgG concentration in the range of 0.2-4.0 nM was obtained with a correlation coefficient of 0.9965. The regression equation was expressed as R = 1.9871C + 2.4616, and a detection limit of 2.9 × 10(-11) M was obtained. The present method was successfully applied for the detection of IgG in human serum. The results indicate that the present protocol is quite promising for the application of CRET in immunoassays. It could also be developed for detection of other antigen-antibody immune complexes by using the corresponding antigens and respective antibodies.

Incorporation of metal-organic framework HKUST-1 into porous polymer monolithic capillary columns to enhance the chromatographic separation of small molecules.

Metal-organic framework (MOF) HKUST-1 nanoparticles have been incorporated into poly(glycidyl methacrylate-co-ethylene dimethacrylate) (HKUST-1-poly(GMA-co-EDMA)) monoliths to afford stationary phases with enhanced chromatographic performance of small molecules in the reversed phase capillary liquid chromatography. The effect of HKUST-1 nanoparticles in the polymerization mixture on the performance of the monolithic column was explored in detail. While the bare poly(GMA-co-EDMA) monolith exhibited poor resolution (Rs<1.0) and low efficiency (800-16,300plates/m), addition of a small amount of HKUST-1 nanoparticles to the polymerization mixture provide high increased resolution (Rs≥1.3) and high efficiency ranged from 16,300 to 44,300plates/m. Chromatographic performance of HKUST-1-poly(GMA-co-EDMA) monolith was demonstrated by separation of various analytes including polycyclic aromatic hydrocarbons, ethylbenzene and styrene, phenols and aromatic acids using a binary polar mobile phase (CH3CN/H2O). The HKUST-1-poly(GMA-co-EDMA) monolith displayed enhanced hydrophobic and π-π interaction characteristics in the reversed phase separation of test analytes compared to the bare poly(GMA-co-EDMA) monolith. The experiment results showed that HKUST-1-poly(GMA-co-EDMA) monoliths are an alternative to enhance the chromatographic separation of small molecules.

Highly sensitive immunoassay of carcinoembryonic antigen by capillary electrophoresis with gold nanoparticles amplified chemiluminescence detection

A noncompetitive immunoassay based on gold nanoparticles (AuNPs) amplified capillary electrophoresis (CE) chemiluminescence (CL) detection was developed for the determination of carcinoembryonic antigen (CEA). In this method, citrate-modified AuNPs were conjugated with horseradish peroxidase (HRP) labeled CEA antibody (Ab*), and incubated with limited amount of CEA antigen. CEA-Ab*-AuNPs complex and excess of Ab*-AuNPs were then separated and quantified by CE with CL detection. Highly sensitive CL detection was achieved by means of p-iodophenol (PIP) enhanced luminol-H2O2-HPR CL reaction and AuNPs amplified. Under the optimal conditions, the CE assay was accomplished within 5min. The linear range for CEA detection was 0.05-20ng/mL with a detection limit of 0.034ng/mL (signal/noise=3), which is three orders magnitude lower than that of without AuNPs amplified. The current method was successfully applied for the quantification of CEA in human serum samples. It was demonstrated that the current CE-CL AuNPs amplified noncompetitive immunoassay was sensitive and highly selective. It may serve as a tool for clinical analysis of CEA to assist in the diagnosis of cancer.

Preparation and characterization of silica monolith modified with bovine serum albumin-gold nanoparticles conjugates and its use as chiral stationary phases for capillary electrochromatography.

This paper describes the development of silica monolith modified with bovine serum albumin-gold nanoparticles (BSA-GNPs) conjugates as chiral stationary phases for capillary electrochromatography (CEC). The bare monolithic silica column was prepared by a sol-gel process and has been modified chemically with 3-mercaptopropyltrimethoxysilane to provide thiol groups, followed by immobilization of gold nanoparticles via the formation of an Au-S bond and modification with BSA as the chiral selector via the nitrogen lone pair of electrons. It has been demonstrated that the monolithic chiral stationary phases can be used for the enantioseparation of a number of phenylthiocarbamyl amino acids (PTC-D/L-AAs) by CEC. Ten pairs of tested amino acids enantiomers were successfully resolved within 18 min under optimized conditions, and the resolution values were in the range of 1.486-2.083. With PTC-D/L-tryptophan used as the probe solute, the influences of applied voltage, organic modifier and buffer pH in mobile phase on apparent retention factor, enantioselectivity and resolution factor were also investigated.

Screening α-glucosidase inhibitor from natural products by capillary electrophoresis with immobilised enzyme onto polymer monolith modified by gold nanoparticles

A novel strategy for screening α-glucosidase inhibitors (AGIs) from natural products by capillary electrophoresis (CE) with an immobilised enzyme microreactor was developed. In this approach, gold nanoparticles (AuNPs) was first covalently attached to surface of the pores of the porous polymer capillary monolith via the formation of an Au-S bond, and α-glucosidase was then simply and stably immobilised onto AuNPs through the strong affinity of gold for amino groups of the enzyme. In order to profiling the activity of the immobilised α-glucosidase, the natural substrate was hydrolyzed by it and the yield of product was determined by CE. The amount of covalently attached α-glucosidase to the monolith was calculated to be about 30.0 μg/mg. The immobilised enzyme exhibited 80% activity after 25 runs, and only lost 7.6% of activity after 6 runs within 31 days. Screening of AGIs present in extracts of natural products by the proposed method was demonstrated.

Noncompetitive immunoassay for carcinoembryonic antigen in human serum by microchip electrophoresis for cancer diagnosis

BACKGROUND Carcinoembryonic antigen (CEA) as one of the most widely used tumor markers is used in the clinical diagnosis of colorectal, pancreatic, gastric, and cervical carcinomas. We describe a microchip electrophoresis (MCE)-based noncompetitive immunoassay technique for assaying CEA in human serum for cancer diagnosis. METHODS Based on a noncompetitive immunoassay format, CEA reacts first with an excess amount of monoclonal antibody (Ab(1)), then free Ab(1) and the bound CEA-Ab(1) complex react with an excess amount of fluorescein isothiocyanate (FITC)-labeled secondary antibody (Ab(2)*) to form CEA-Ab(1)-Ab(2)* and Ab(1)-Ab(2)* complexes. Finally, the free Ab(2)* and bound CEA-Ab(1)-Ab(2)*, Ab(1)-Ab(2)* complexes were separated and detected by MCE coupling laser-induced fluorescence (LIF), and CEA was quantified by measuring the fluorescence intensity of CEA-Ab(1)-Ab(2)*. RESULTS The linear range for CEA was 60pg/ml-8ng/ml with a correlation coefficient of 0.9992 and a detection limit of 45.7pg/ml. The immunocomplex including CEA-Ab(1)-Ab(2)*, Ab(1)-Ab(2)*and free Ab(2)* was separated within 80s. The amounts of CEA in normal person serum were found to be in the range of 2.9-5.1microg/l, and seven cancer patients serum were analyzed, and CEA levels range from 79.6-270.1ng/ml. CONCLUSIONS This method may become a useful tool for rapid analysis of CEA and other tumor markers in biomedical analysis and clinical diagnosis.

Synthesis of magnetic porous γ-Fe2O3/C@HKUST-1 composites for efficient removal of dyes and heavy metal ions from aqueous solution

A novel and inexpensive approach was adopted to develop magnetic porous γ-Fe2O3/C@HKUST-1 composites for the adsorption of dyes and heavy metal ions from aqueous solution. The γ-Fe2O3/C with unique functional groups present such as –OH and –NH2 was used as the support to directly grow HKUST-1 by a stepwise liquid-phase epitaxy process. The crystallographic, morphology, and magnetic properties as well as porosity of the as-synthesized γ-Fe2O3/C @HKUST-1 composites were carefully studied by XRD, SEM, TEM, XPS, TGA, and BET. The results indicated that the BET surface area, micropore volume, and saturation magnetization of the γ-Fe2O3/C @HKUST-1 are 993.4 m2 g−1, 0.69 cm3 g−1, and 12.6 emu g−1, respectively. In addition, a uniform distribution of ultrafine γ-Fe2O3 nanoparticles with an average diameter of 2–3 nm was observed in the γ-Fe2O3/C@HKUST-1 composites. Our results showed that methylene blue (MB) and Cr(VI) (used as a model for typical dye pollutants and heavy metal ions) are effectively removed from aqueous solutions by γ-Fe2O3/C@HKUST-1. The maximum adsorption capacities were 370.2 and 101.4 mg g−1 of adsorbent for MB and Cr(VI), respectively. Moreover, a removal efficiency of about 90% was retained after five cycles of consecutive adsorption–desorption. The adsorption kinetics data were well described by a pseudo-second-order model (R2 > 0.99), and equilibrium data were well fitted to the Langmuir isotherm model (R2 > 0.99). Finally, our results suggested that the γ-Fe2O3/C@HKUST-1 composites have a great potential to be employed for treatment of wastewater containing MB and Cr(VI).

Determination of intracellular sulphydryl compounds by microchip electrophoresis with selective chemiluminescence detection.

An analytical method based on microchip electrophoresis (MCE) and chemiluminescence detection (CL) was developed for the determination of intracellular sulphydryl compounds. Cell injection/loading, cytolysis, electrophoretic separation, and CL detection were integrated onto a simple cross-microfluidic chip. Selective CL detection of sulphydryl compounds was achieved by deploying the luminol-Na(2)S(2)O(8) reaction. Under the CL conditions selected, many endogenous compounds in biological systems such as amino acids, biogenic amines, peptides and proteins did not produce any CL signal, which further ensured a high selectivity of the proposed MCE-CL assays. Sulphydryl compounds including cysteine (Cys), glutathione (GSH), and hemoglobin (Hb) were selected as the test compounds. The MCE separation was completed within 120s. The detection limits were estimated to be 7amol for Cys, 32amol for GSH and 69amol for Hb, respectively. The present method was applied to analyze individual red blood cells collected from both healthy subjects and cancer patients. It was found that the average intracellular contents of Cys, GSH and Hb were in the ranges of 26-43amol/cell, 128-323amol/cell and 522-667amol/cell, respectively for cancer patients, compared to 579-609amol Hb/cell and not detectable Cys and GSH for healthy subjects.

From metal–organic frameworks to magnetic nanostructured porous carbon composites: towards highly efficient dye removal and degradation

A magnetic nanostructured porous carbon material (γ-Fe2O3/C) was easily synthesized using a microwave-enhanced high-temperature ionothermal method with an iron terephthalate metal–organic framework-MIL-53(Fe), as a template. The structure, morphology, magnetic properties, and porosity of γ-Fe2O3/C were characterized by powder X-ray diffraction, Raman spectroscopy, X-ray photoelectron spectroscopy, scanning electron microscopy, high-resolution transmission electron microscopy, vibrating sample magnetometry, and Brunauer–Emmett–Teller surface area analysis. The obtained porous carbon materials (γ-Fe2O3/C), possessed a high specific surface area (397.2 m2 g−1) and pore volume (0.495 cm2 g−1). The adsorption properties were tested by removal of malachite green (MG) from an aqueous solution. After reaching adsorption equilibrium, the maximum adsorption capacity was 499 mg g−1 at 30 °C, and reached 863 mg g−1 at 60 °C. The excellent magnetism (20.10 emu g−1) provided an ideal magnetic-separation performance. Analysis of the sorption kinetics and isotherms showed that these sorption processes were better fitted to the pseudo-second-order and Langmuir equations than pseudo-first-order and Freundlich equations. Various thermodynamic parameters, such as ΔGθ, ΔSθ, and ΔHθ, were also calculated, and indicated that the present system was spontaneous and endothermic. It was further demonstrated that γ-Fe2O3 showed powerful photocatalytic activity for the degradation of MG under sunlight in the presence of H2O2.