OBJECTIVETo improve the key technology of immunesensors in immobilizing bio-sensitive element and keeping its bioactivity, an enzyme immunosensor based on chitosan-SiO(2) (CS-Sio(2)) hybrid membrane was fabricated. To estimate the new immunosensor Vibrio parahaemolyticus which was the main pathogens of aquatic products.

METHODSA CS-SiO(2) hybrid membrane was prepared using sol-gel method. The enzyme immunosensor was fabricated by coating the membrane and horseradish peroxidase labeled Vibrio parahaemolyticus antibody (HRP-anti-VP) on the surface of four-channel screen-printed carbon electrode. The immunosensor was characterized by cyclic voltammetry. Vibrio parahaemolyticus could be detected according to the decrease percentage (DP) of peak current before and after immune response, while cyclic voltammetry was used as an electrochemical mean to detect the products of the enzymatic reaction. Seven kinds of bacteria, like Vibrio alginolyticus, were selected for specific experiments.

RESULTSBy studying the infrared spectrum of three kinds of films, the CS-SiO(2) hybrid membrane was prepared and HRP-anti-VP was fixed in the hybrid membrane. Under the optimum conditions of immunoreaction and electrochemical detection, the DP of peak current before and after immune response showed a linear relation with lgC in the range of 10(4) - 10(9) cfu/ml, while the linear regression equation was: DP = 6.5 lgC-3.319, the correlation coefficient was 0.9958 and the detection limit was 6.9 x 10(3) cfu/ml (S/N = 3). The immunosensor possessed acceptable specificity, reproducibility (RSD < 6%), stability (the amperometric response was 95% of the initial response after a week) and accuracy (96.7% of the results obtained by the immunosensor were in agreement with those obtained by GB/T 4789.7-2003).

CONCLUSIONThe enzyme immunosensor based on CS-SiO(2) hybrid membrane gave a good performance in rapid detection of Vibrio parahaemolyticus.

Biosensing Techniques ; instrumentation ; methods ; Electrochemical Techniques ; instrumentation ; methods ; Equipment Design ; Immunoenzyme Techniques ; instrumentation ; methods ; Silicon Dioxide ; Vibrio parahaemolyticus ; isolation & purification

p-Nitrophenylphosphate (PNPP) is usually employed as the substrate for enzyme-linked immunosorbent assays. p-Nitrophenol (PNP), the product of PNPP, with the catalyst alkaline phosphatase (ALP), will passivate an electrode, which limits applications in electrochemical analysis. A novel anti-passivation ink used in the preparation of a graphene/ionic liquid/chitosan composited (rGO/IL/Chi) electrode is proposed to solve the problem. The anti-passivation electrode was fabricated by directly writing the graphene-ionic liquid-chitosan composite on a single-side conductive gold strip. A glassy carbon electrode, a screen-printed electrode, and a graphene-chitosan composite-modified screen-printed electrode were investigated for comparison. Scanning electron microscopy was used to characterize the surface structure of the four different electrodes and cyclic voltammetry was carried out to compare their performance. The results showed that the rGO/IL/Chi electrode had the best performance according to its low peak potential and large peak current. Amperometric responses of the different electrodes to PNP proved that only the rGO/IL/Chi electrode was capable of anti-passivation. The detection of cardiac troponin I was used as a test example for electrochemical immunoassay. Differential pulse voltammetry was performed to detect cardiac troponin I and obtain a calibration curve. The limit of detection was 0.05 ng/ml.
Electrochemical Techniques/methods* ; Electrodes ; Graphite ; Immunoassay/methods* ; Ink ; Microscopy, Electron, Scanning ; Troponin I/blood*

To establish an electrochemiluminescence immunoassay (ECLIA) for the detection of influenza virus B (Flu B), we biotinylated the Flu B monoclonal antibody (mAb),labeled the paired Ab with ruthenium and used the streptavidin-coated microparticles in this study. The ECL intensity was linear with the concentrations of antigen of Flu B in the range of 0.55 microg/mL to 17.5 microg/mL, with a detection limit of 0.55 microg/mL (S/N= 3). The precision,sensitivity and specificity were evaluated. The established ECLIA for Flu B antigen in this study was specific, sensitive and precise. It may provide a convenient and rapid technique for clinical detection of Flu B.
Antibodies, Viral ; analysis ; Electrochemical Techniques ; Immunoassay ; methods ; Influenza B virus ; isolation & purification ; Luminescence

OBJECTIVETo analyze the clinical performance of free prostate-specific antigen (fPSA) detection by ECLIA method, and evaluate whether ECLIA is suitable for clinical use.

METHODS341 samples were collected and tested prostate-specific antibodies with CMIA and ECLIA methods. These samples contain: 97 samples with abnormal high PSA value tested by CMIA method, and 244 normal PSA samples. Use CMIA as the reference method, and detect fPSA, tPSA levels, and the ratio of fPSA/tPSA. Analyze the testing results with statistical methods.

RESULTSCompared with CMIA, correlation coefficent of ECLIA fPSA detection is 0.99; correlation coefficent of f/tPSA ratio detection is 0.96; the sensitivity, specificity of ECLIA f/tPSA ratio detection are 85.71%, 92.6% respectively, the agreement rate with ECLIA is 87.4%. No cross reaction with bilirubin, lipohemia, hemolysis, RF, CEA, AFP, CA125, CA153, CA199 were found in the tests.

CONCLUSIONThe ECLIA method for free prostate-specific antigen detection showed good clinical performance; and is suitable for clinical use.

Electrochemical Techniques ; methods ; Humans ; Male ; Prostate-Specific Antigen ; blood ; Prostatic Neoplasms ; blood ; diagnosis ; Sensitivity and Specificity

In this study, we investigated reductive degradation of nitroaromatic antibiotic chloramphenicol to non-effective antibacterial amine product in fed-batch biocatalyzed electrolysis systems (BES) (applied voltage was 0.5 V) under low temperature (12 +/- 2 degrees C). The ohm resistance of the whole BES reactor increased when the phosphate buffer solution concentrations decreased. Efficiencies (ErCAP) of chloramphenicol reduction with biocathode (PBS, 25 mmol/L) in presence of glucose was (86.3 +/- 1.69)% within 24 h and sludge fermentation liquor was (74.1 +/-1.44)% within 24 h. While the ErCAP of abiotic cathode under the same condition was only (57.9 +/- 1.94)% within 24 h. It suggested that biocathode could be a promising technology for reductive biodegradation of nitroaromatic antibiotics-containing wastewater in areas with relatively low annual mean temperature.
Bioreactors ; Chloramphenicol ; metabolism ; Electrochemical Techniques ; instrumentation ; methods ; Electrodes ; Fermentation ; Oxidation-Reduction

This study was to develop a new method for detecting circulating tumor cells (CTCs) with high sensitivity and specificity, therefore to detect the colorectal cancer as early as possible for improving the detection rate of the disease. To this end, we prepared some micro-column structure microchips modified with graphite oxide-streptavidin (GO-SA) on the surface of microchips, further coupled with a broad-spectrum primary antibody (antibody1, Ab₁), anti-epithelial cell adhesion molecule (anti-EpCAM) monoclonal antibody to capture CTCs. Besides, carboxylated multi-walled carbon nanotubes (MWCNTs-COOH) were coupled with colorectal cancer related antibody as specific antibody 2 (Ab₂) to prepare complex. The sandwich structure consisting of Ab₁-CTCs-Ab₂ was constructed by the microchip for capturing CTCs. And the electrochemical workstation was used to detect and verify its high sensitivity and specificity. Results showed that the combination of immunosensor and micro-nano technology has greatly improved the detection sensitivity and specificity of the immunosensor. And we also verified the feasibility of the immunosensor for clinical blood sample detection, and successfully recognitized detection and quantization of CTCs in peripheral blood of colorectal cancer patients by this immunosensor. In conclusion, the super sandwich immunosensor based on micro-nano technology provides a new way for the detection of CTCs, which has potential application value in clinical diagnosis and real-time monitoring of disease.
Humans ; Nanotubes, Carbon/chemistry* ; Neoplastic Cells, Circulating/pathology* ; Biosensing Techniques ; Immunoassay/methods* ; Antibodies ; Colorectal Neoplasms/diagnosis* ; Electrochemical Techniques/methods* ; Gold/chemistry*

OBJECTIVETo find the potential interference factors for the detection of NT-proBNP and BNP in patients with chronic heart failure.

METHODSEP15-A2 issued by Clinical and Laboratory Standards Institute (CLSI) was employed to compare the precision and accuracy of commercial NT-proBNP and BNP analyzer electrochemiluminescence immunoassay system Cobas E601 and chemiluminescence system ADVIA Centaur. Moreover, NT-proBNP and BNP were detected in different time interval and in different interfered sampling conditions (haematolysis, choloplania, lipemia). NT-proBNP and BNP of 203 patients with heart failure or heart failure complicated with acute cerebral infarction were analyzed to find the deviation caused by patients' endogenous factors.

RESULTSThe precision and accuracy were comparable for NT-proBNP and BNP detection using Cobas E601 and ADVIA Centaur (total-CV below 2.9% and 3.5%, the deviation from definite value below 2.38% and 3.91%). The most suitable sample type for NT-proBNP and BNP detection was serum and EDTA-anticoagulant plasma. The detection results of NT-proBNP and BNP were comparable for at least 120 min post sampling and not affected by Hb (2 g/L), DB (428 µmol/L) and chyle (2000 FIU). NT-proBNP was significantly higher in heart failure patients complicated with cerebral infarction (P = 0.003) than in heart failure patients. BNP was significantly higher in heart failure grade III patients complicated with cerebral infarction (P < 0.01).

CONCLUSIONSCobas E601 and ADVIA Centaur supplied satisfactory detection of NT-proBNP and BNP in patients with chronic heart failure with strong anti-interference capacity. The diagnostic value of NT-proBNP and BNP for chronic heart failure should be analyzed objectively in the presence of complicating diseases.

Electrochemical Techniques ; methods ; Heart Failure ; blood ; diagnosis ; Humans ; Immunoassay ; methods ; Luminescent Measurements ; methods ; Natriuretic Peptide, Brain ; blood ; Peptide Fragments ; blood ; Sensitivity and Specificity ; Specimen Handling ; methods ; standards

We studied the interaction between proteins and polystyrene-polyacrylic acid (PS-PAA) spherical polyelectrolyte brushes with different polyacrylic acid (PAA) chain lengths, including the physical adsorption and chemical adsorption in PBS buffer. Results showed that the amount of bovine serum albumin (BSA) physically adsorbed on PS-PAA spherical polyelectrolyte brushes decreased to a minimum of 33 microg/mg whereas the amount of streptavidin (SA) chemically adsorbed increased with the increase of chain length and carboxyl quantity. The biotin binding capacity of streptavidin chemically adsorbed on PS-PAA spherical polyelectrolyte brushes was roughly evaluated via enzyme competitive inhibition.
Acrylic Resins ; chemistry ; Adsorption ; Biosensing Techniques ; Electrochemical Techniques ; methods ; Electrolytes ; chemistry ; Polyamines ; Polymers ; chemistry ; Polystyrenes ; chemistry ; Proteins ; chemistry ; Serum Albumin, Bovine ; chemistry ; Surface Properties

A novel type of carbon nanotube-coated Au nanoparticle and [bmim]BF4 composite modified glassy carbon electrode was fabricated by a layer-by-layer self-assembly technique. The electrochemical performance of acetaminophen (ACOP) on the modified electrode was investigated by cyclic voltammetry. The Nafion/GNPs/RTIL/MWNTs/GC electrode showed an excellent electrocatalytic activity for the oxidation of ACOP and accelerated electron transfer between the electrode and ACOP. For ACOP, the reversible electrochemical process was observed on the Nafion/GNPs/RTIL/MWNTs/GC electrode, while irreversible electrochemical process occurred on the GC electrode. For the Nafion/GNPs/RTIL/MWNTs/GC electrode, the anodic peak potential of ACOP was moved from 0.562 V to 0.413 V, with a potential drop of 149 mV. At the same time, the reduction peak potential was 0.384 V, and the potential difference was only 29 mV. It was shown that the modified electrode possessed higher electrocatalytic activity and more sensitive effect for the detection of ACOP than both MWNTs/GC electrode and GC electrode. The effects of the different experimental conditions on the electrochemical behaviors of ACOP were explored. Under the optimum conditions of preparation and experimental, the linear calibration curves of ACOP were obtained in a wide range of 2 x 10(-1) to 4.0 x 10(-4) mol x L(-1) with a correlation coefficient 0.999 2 and a detection limit of 2.6 x 10(-8) mol x L(-1) (the ratio of signal to noise, 3:1). The recovery rate was 97.9%-100.8%. This method can be used to determine ACOP in paracetamol tablets with satisfactory results.
Acetaminophen ; analysis ; Analgesics, Non-Narcotic ; analysis ; Antipyretics ; analysis ; Electrochemical Techniques ; methods ; Electrochemistry ; methods ; Electrodes ; Electron Transport ; Gold ; chemistry ; Nanotubes, Carbon ; chemistry ; Oxidation-Reduction ; Reproducibility of Results

Animals ; Electrochemical Techniques ; methods ; Gene Silencing ; Genes, Reporter ; Humans ; MicroRNAs ; blood ; genetics ; metabolism ; physiology ; Microarray Analysis ; RNA, Messenger ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction