ELECTROCHEMICAL CYSTEINE DETERMINATION IN SERUM SAMPLES BY Hg THIN FILM SENSOR

Cysteine is a nonessential aminoacid, meaning that cysteine can be made in the human body. It is one of the few amino acids that contain sulfur. This allows cysteine to bond in a special way and maintain the structures of proteins in the body. Cysteine strengthens the protective lining of the stomach and intestines, which may help prevent damage caused by aspirin and similar drugs. In addition, cysteine may play an important role in the communication between immune system cells.

In this study, glassy carbon electrodes modified with mercury (Hg) were used as working electrode. Mercury thin film on glassy carbon electrode was deposited by holding the electrode potential at ?0.7 V; the measurement period for the coating process was 2 minutes. pH and temperature effects on the electrode response were carried out by working at different pHs and temperatures. The calibration graph for cysteine was drawn in the range of 5–120 μM cysteine. Repeatability and interferences studies were investigated. GSH had an interference effect of about 13% of cysteine response. Finally, the sensor was applied to real samples for cysteine determination and the method was validated by Ellman's reagent.     

Kinetic Analysis of Glucoamylase-Catalyzed Hydrolysis of Starch Granules from Various Botanical Sources

The kinetics of glucoamylase-catalyzed hydrolysis of starch granules from six different botanical sources (rice, wheat, maize, cassava, sweet potato, and potato) was studied by the use of an electrochemical glucose sensor. A higher rate of hydrolysis was obtained as a smaller size of starch granules was used. The adsorbed amount of glucoamylase on the granule surface per unit area did not vary very much with the type of starch granules examined, while the catalytic constants of the adsorbed enzyme (k ₀) were determined to be 23.3±4.4, 14.8±6.0, 6.2±1.8, 7.1±4.1, 4.6±3.0, and 1.6±0.6 s^?1 for rice, wheat, maize, cassava, sweet potato, and potato respectively, showing that k ₀ was largely influenced by the type of starch granules. A comparison of the k ₀-values in relation to the crystalline structure of the starch granules suggested that k ₀ increases as the crystalline structure becomes dense.     

Nanostructure shape effects on response of plasmonic aptamer sensors

A localized surface plasmon resonance (LSPR) sensor surface was fabricated by the deposition of gold nanorods on a glass substrate and subsequent immobilization of the DNA aptamer, which specifically bind to thrombin. This LSPR aptamer sensor showed a response of 6‐nm λ_max shift for protein binding with the detection limit of at least 10 pM, indicating one of the highest sensitivities achieved for thrombin detection by optical extinction LSPR. We also tested the LSPR sensor fabricated using gold bipyramid, which showed higher refractive index sensitivity than the gold nanorods, but the overall response of gold bipyramid sensor appears to be 25% less than that of the gold nanorod substrate, despite the approximately twofold higher refractive index sensitivity. XPS analysis showed that this is due to the low surface density of aptamers on the gold bipyramid compared with gold nanorods. The low surface density of the aptamers on the gold bipyramid surface may be due to the effect of shape of the nanostructure on the kinetics of aptamer monolayer formation. The small size of aptamers relative to other bioreceptors is the key to achieving high sensitivity by biosensors on the basis of LSPR, demonstrated here for protein binding. The generality of aptamer sensors for protein detection using gold nanorod and gold nanobipyramid substrates is anticipated to have a large impact in the important development of sensors toward biomarkers, environmental toxins, and warfare agents. Copyright © 2013 John Wiley & Sons, Ltd.     

Direct electrochemistry and electrocatalysis of hemoglobin on polypyrrole-Fe3O4/dodecyltrimethylammonium bromide-modified carbon paste electrode and its biosensing for hydrogen peroxide

Abstract

A novel hydrogen peroxide (H₂O₂) biosensor was successfully constructed, based on the immobilization of hemoglobin (Hb) on polypyrrole (PPy)-Fe₃O₄ and dodecyltrimethylammonium bromide (DTAB) composite ?lm-modified carbon paste electrodes (CPE). The PPy-Fe₃O₄ composites were synthesized in the suspension solution of Fe₃O₄ nanoparticles via in situ chemical oxidative polymerization under the direction of cationic surfactant cetyl trimethyl ammonium bromide. Spectroscopic and electrochemical examinations illustrated that the PPy-Fe₃O₄/DTAB composites were a biocompatible matrix for immobilizing Hb, which revealed high chemical stability and excellent biocompatibility. The thermodynamic, dynamic, and catalytic performance of the biosensor were analysed using cyclic voltammetry (CV). The results indicated that the PPy-Fe₃O₄/Hb/DTAB/CPE exhibited excellent electrocatalytic activity in the reduction of H₂O₂ with a high sensitivity (104 μA mM^{? 1}). The catalytic reduction currents of H₂O₂ were linearly related to H₂O₂ concentration in the range from 2.5 μM to 60 μM with a detection limit of 0.8 μM (S/N = 3). With such superior characteristics, this biosensor for H₂O₂ can be potentially applied in determination of other reactive oxygen species as well. These results indicated that PPy-Fe₃O₄/DTAB composites are a promising matrix for bioactive molecule immobilization.     

Biosensor for acrylamide based on an ion-selective electrode using whole cells of Pseudomonas aeruginosa containing amidase activity

The aim of this work was to develop a biosensor for toxic amides using whole cells of Pseudomonas. aeruginosa containing amidase activity, which catalyses the hydrolysis of amides such as acrylamide producing ammonia and the corresponding organic acid. Whole cells immobilized in several types of membrane in the presence of glutaraldehyde and an ammonium ion-selective electrode, were used for biosensor development. This biosensor exhibited a linear response in the range of 0.1–4.0×10^?3 M of acrylamide, a detection limit of 4.48×10^?5 M acrylamide, a response time of 55 s, a sensitivity of 58.99 mV mM^?1 of acrylamide and a maximum t_1/2 of 54 days. The selectivity of this biosensor towards other amides was investigated, which revealed that it cross-reacted with acetamide and formamide, but no activity was detected with phenylacetamide, p-nitrophenylacetamide and acetanilide. It was successfully used for quantification of acrylamide in real industrial effluents and recovery experiments were carried out which revealed an average substrate recovery of 93.3%. The biosensor is cheap since whole cells of P. aeruginosa can be used as source of amidase activity.     

DNA entrapped polypyrrole-polyvinyl sulfonate film for application to electrochemical biosensor

Double-stranded calf thymus (dsCT)-DNA was electrochemically entrapped into polypyrrole-polyvinyl sulfonate (PPy-PVS) films deposited onto indium tin oxide (ITO) coated glass plates. These dsCT-DNA entrapped PPy-PVS/ITO films were characterized using cyclic voltammetry, UV-visible, Fourier transform infrared (FT-IR), scanning tunneling microscopy (STM), and electrochemical impedance measurements. Attempts made to use these dsCT-DNA entrapped PPy-PVS/ITO films for detection of 2-aminoanthracene (0.001-6.0 ppm) and 3-chlorophenol (0.01-55.0 ppm) revealed a response time of 30s and a shelf life of approximately 25 weeks when stored under desiccated conditions at 25 degrees C. The addition of salts such as Ca(2+) (250 ppm), Mg(2+) (200 ppm), Cl(-) (1560 ppm), and Na(+) (150 ppm) ions contained in water does not affect the observed amperometric response of the disposable dsCT-DNA entrapped PPy-PVS film-based electrochemical biosensor.     

A nucleic acid biosensor for the detection of a short sequence related to the hepatitis B virus using bis(benzimidazole)cadmium(II) dinitrate as an electrochemical indicator

A novel hybridization indicator, bis(benzimidazole)cadmium(II) dinitrate (Cd(bzim)(2)(NO(3))(2)), was utilized to develop an electrochemical DNA biosensor for the detection of a short DNA sequence related to the hepatitis B virus (HBV). The sensor relies on the immobilization and hybridization of the 21-mer single-stranded oligonucleotide from the HBV long repeat at the glassy carbon electrode (GCE). The hybridization between the probe and its complementary sequence as the target was studied by enhancement of the peak of the Cd(bzim)(2)(2+) indicator using cyclic voltammetry (CV) and differential pulse voltammetry (DPV). Numerous factors affecting the probe immobilization, target hybridization, and indicator binding reactions were optimized to maximize the sensitivity and speed of the assay time. With this approach, a sequence of the HBV could be quantified over the range from 1.49x10(-7)M to 1.06x10(-6)M, with a linear correlation of r=0.9973 and a detection limit of 8.4x10(-8)M. The Cd(bzim)(2)(2+) signal observed from the probe sequence before and after hybridization with a four-base mismatch containing sequence was lower than that observed after hybridization with a complementary sequence, showing good selectivity. These results demonstrate that the Cd(bzim)(2)(2+) indicator provides great promise for the rapid and specific measurement of the target DNA.     

基于CRISPR/Cas体系的生物传感器在病原体核酸检测方面的应用

成簇规律间隔短回文重复序列/成簇规律间隔短回文重复序列相关蛋白 (Clustered regularly interspaced short palindromic repeats/CRISPR-associated protein,CRISPR/Cas) 因高效的靶向结合和可编程性,已被开发为一种精准、高效、低价和高灵敏度的核酸检测工具。目前基于CRISPR-Cas体系的生物传感器在病原体核酸检测方面显示出了优良的性能,受到了人们的广泛关注,这种新型的病原体核酸检测有望替代传统的病原体检测方法。文中就基于CRISPR/Cas体系的生物传感器在病原体核酸检测中的最新研究进展进行综述。     

An amperometric biosensor based on horseradish peroxidase immobilized onto maize tassel-multi-walled carbon nanotubes modified glassy carbon electrode for determination of heavy metal ions in aqueous solution

A biosensor for trace metal ions based on horseradish peroxidase (HRP) immobilized on maize tassel-multiwalled carbon nanotube (MT-MWCNT) through electrostatic interactions is described herein. The biosensor was characterized using Fourier transform infrared (FTIR), UV–vis spectrometry, voltammetric and amperometric methods. The FTIR and UV–vis results inferred that HRP was not denatured during its immobilization on MT-MWCNT composite. The biosensing principle was based on the determination of the cathodic responses of the immobilized HRP to H₂O₂, before and after incubation in trace metal standard solutions. Under optimum conditions, the inhibition rates of trace metals were proportional to their concentrations in the range of 0.092–0.55 mg L⁻¹, 0.068–2 mg L⁻¹ for Pb²⁺ and Cu²⁺ respectively. The limits of detection were 2.5 μg L⁻¹ for Pb²⁺ and 4.2 μg L⁻¹ for Cu²⁺. Representative Dixon and Cornish-Bowden plots were used to deduce the mode of inhibition induced by the trace metal ions. The inhibition was reversible and mixed for both metal ions. Furthermore, the biosensor showed good stability, selectivity, repeatability and reproducibility.     

Amperometric bioelectrode for specific human immunoglobulin G determination: optimization of the method to diagnose American trypanosomiasis

Bioelectrodes to detect immunoglobulin G (IgG) antibodies occurring in sera of patients suffering from American trypanosomiasis were assembled. The device consisted of a gold electrode modified with a thiol sensitized with parasite proteins. The assemblage was accomplished by adsorbing IgG antibodies from confirmed infected patients followed by adsorption of anti-human IgG labeled with a redox enzyme. The appliance was used as a working electrode in a three-electrode cell containing a soluble charge-transfer mediator, also behaving as enzyme cosubstrate. The method is based on the measurement of the catalytic current after addition of the enzyme substrate, occurring when a positive serum is used to build up the biosensor. The discrimination efficiency between positive and negative sera was 100% for the samples studied. A 0.9525 correlation coefficient was obtained for results acquired by using this approach and one commercial diagnostic kit. The reproducibility, evaluated by the percentage coefficient of variation, varied between 7 and 20%. The sensitivity was 12.4 ng mL(-1) IgG, which is in the same order as that obtained with the commercial kit. Stability of the device was studied for a 7-day period and the results showed no significant change (p = 0.218). Leishmaniasic sera showed cross-reactivity when total parasite homogenate was used as antigen.