Kosakonia radicincitans and Cryptococcus laurentii controlled Penicillium expansum rot and decreased patulin production at 4 and 25 °C.

In the present work, we evaluated the effects of a mixture of biocontrol agents against two toxigenic strains of Penicillium expansum isolated in Argentine Patagonia from pome fruits. The two strains, INTA-5 and INTA-10, were previusly selected among ten strains coming from the Alto Valle (Rio Negro-Argentina) for their high production of patulin. For the biocontrol, Kosakonia radicincitans, Cryptococcus laurentii, and Rhodosporidium fluviale were tested in vitro experiments on Potato Dextrose Agar (PDA) dishes against the INTA-5 and INTA-10 strains. The bacterium K. radicincitans and the yeast C. laurentii were selected to be used in a mixture due to their capacity to control the fungus and reduce the mycotoxin severely. In vitro assays with the mixture showed a high antagonism against P. expansum INTA-5 and INTA-10, at 21 d of incubation at 25 °C and a patulin reduction of 98%. The mixture of microorganisms was also effective in apples stored at 25 °C for 10 d and 4 °C for 30 d. At cold storage, the mixture controlled moderately the development of rot and decreased patulin concentration. At 25 °C, the pathogen's optimal growth temperature, the mixture of Biological Control Agent (BCAs) assured both the control of rot and decrease of patulin concentration. The combination of two microorganisms, with different requirements and abilities, resulted in a mix with a strong antagonism against P. expansum with the capability to decrease the patulin concentration. Treatment with the selected mixture could be a good option for controlling strains with different behaviours and in different environmental conditions.

First steps in the formulation of praziquantel nanosuspensions for pharmaceutical applications.

Praziquantel (PZQ), a broad spectrum anthelmintic drug, cannot be found in acceptable dosage forms for elderly patients, paediatric patients, and for veterinary use. In fact, very little has been done up to now in the formulation of liquid dosage forms, being they always formulated for parenteral administration. To beat this important challenge, it was accomplished a comprehensive analysis of the influence of two elementary physicochemical aspects, i.e. surface thermodynamic and electrokinetic properties, on the colloidal stability of PZQ nanosuspensions. The hydrophobic character of the drug, intensely determining the flocculation curves, was confirmed by the thermodynamic characterization. The electrophoretic characterization, in combination with the sedimentation and relative absorbance versus time curves, highlighted that the electrical double layer thickness and the surface charge can play an essential role in the stability of the pharmaceutical colloid. Finally, it was demonstrated that controlling the pH values and the incorporation of electrolytes can help in formulating PZQ aqueous nanosuspensions with appropriate stability and redispersibility behaviours for pharmaceutical use.

Serological diagnosis of Toxoplasmosis disease using a fluorescent immunosensor with chitosan-ZnO-nanoparticles.

This article describes a microfluidic LIF immunosensor for the quantitative determination of anti-Toxoplasma gondii IgG (anti-T. gondii) specific antibodies. The serological detection of these antibodies plays a crucial role in the clinical diagnosis of toxoplasmosis. Zinc oxide nanoparticles (ZnO-NPs) obtained by wet chemical procedure were covered with chitosan and then used to conjugate T-gondii antigens into the central microfluidic channel. Serum samples containing anti-T-gondii IgG antibodies were injected into the immunosensor where they interact immunologically with T. gondii antigens. Bound antibodies were quantified by the addition of anti-IgG antibodies labeled whit alkaline phosphatase (ALP). ALP enzymatically converts the non-fluorescent 4-methylumbelliferyl phosphate (4-MUP) to soluble fluorescent methylumbelliferone that was measured using excitation at 355 nm and emission at 440 nm. The relative fluorescent response of methylumbelliferone is proportional to the concentration of anti-T. gondii IgG antibodies. The coefficients of variation are less than 4.73% for within-day assays and less than 6.34% for between-day assays. Results acquired by LIF immunosensor agree with those obtained by enzyme-linked immunosorbent assay method, suggesting that the designed sensor represents a promising tool for the quantitative determination of anti-T. gondii IgG antibodies of clinical samples.

Novel Electrochemical Paper-Based Immunocapture Assay for the Quantitative Determination of Ethinylestradiol in Water Samples.

We report a novel and innovative electrochemical paper-based immunocapture assay (EPIA) to address the need for ultrasensitive detection of emerging pollutants without regulatory status and whose effects on environment and human health are not completely yet understood. In particular, we present the application of this system toward highly sensitive detection of the emerging pollutant ethinyl estradiol (EE2). The EPIA approach is based on the use of paper microzones modified with silica nanoparticles (SNs) and anti-EE2 specific antibodies for capture and preconcentration of EE2 from river water samples. After the preconcentration procedure, the paper microzones are placed onto a screen-printed carbon electrode modified with electrochemically reduced graphene (RG). The bound EE2 is subsequently desorbed adding a diluted solution of sulfuric acid on the paper microzones. Finally, recovered EE2 is electrochemically detected by OSWV. The proposed novel methodology showed an appropriate LOD and linear range for the quantification of EE2 for water samples with different origins. The nonsophisticated equipment required, the adequate recovery values obtained (from 97% to 104%, with a RSD less than 4.9%), and the appropriate LOD and linear range value (0.1 ng L-1 and 0.5-120 ng L-1, respectively) achieved by our immunocapture sensor present significant analytical figures of merit, particularly when the routine quantification of EE2 is considered. In addition, our system was based on electrochemical paper-based technology, which allows obtainment of portable, easy-to-use, inexpensive, and disposable devices. The EPIA can also serve as a general-purpose immunoassay platform applicable to quantitation of other drugs and emerging pollutants in environmental samples.

Assessment of ochratoxin A occurrence in Argentine red wines using a novel sensitive quechers-solid phase extraction approach prior to ultra high performance liquid chromatography-tandem mass spectrometry methodology.

BACKGROUND: The assessment of ochratoxin A (OTA) in wine is relevant for food safety and its continuous control allows to reduce the risk of intake. Thus, a novel sensitive QuEChERS-SPE (Quick, Easy, Cheap, Effective, Rugged and Safe - Solid Phase Extraction) pretreatment prior to liquid chromatography coupled to tandem mass spectrometry was developed for the determination of OTA in red wine samples from different grape-growing regions in Argentine. RESULTS: A sensitive methodology was achieved and thus the limits of detection and quantification were 0.02 and 0.05 µg L-1 , respectively. Recoveries ranged from 89.0% to 105.3%. The method was applied to 136 red wine samples (Argentina's flagship varieties: Malbec and Cabernet Sauvignon) from ten grape-growing regions, during vintages 2013-2015. Although all of the samples investigated were contaminated with OTA (concentrations ranged from 0.02 to 0.98 µg L-1 ), the levels detected were lower than the maximum allowable concentration limit of 2.0 µg L-1 established by international regulations. CONCLUSION: The methodology proposed is suitable for reliable OTA analysis in red wines. Similarly, the values obtained from the samples analyzed were in accordance with the current regulations and, as a consequence, preventive actions to reduce this mycotoxin incidence can be undertaken. © 2016 Society of Chemical Industry.

A green alternative method for analysis of ivermectin and moxidectin in environmental water samples using automatized preconcentration previous MEEKC.

Antiparasitic drugs derived from macrocyclic lactones (MLs) are widely used in livestock activities around the world. An increasing concern for local authorities is the environmental pollution as a consequence of veterinary drugs widely used in rural areas. The purpose of environmental analysis is to monitor low levels of pollutant analytes in a large number of samples. Also, due to the lipophilic characteristic of these lactones, long-chain solvents are usually required for performing sample treatment before and during the analysis. Therefore, sensitive, specific, robust, and environmentally friendly analytical methods are still required. In this paper, a new automatized preconcentration methodology followed by microemulsion electrokinetic chromatography analysis was developed for the simultaneous separation and determination of the most used MLs, ivermectin (IVM) and moxidectin (MXD) in environmental water. XAD-4 resin was employed as an adsorbent for the preconcentration process and ethanol was used as the eluent. In contrast to traditional analysis for IVM and MXD, in this methodology nonpolluting solvents were involved during the whole process and therefore, it could be considered as a contribution to green analytical chemistry. Under optimal experimental conditions, LOD obtained for IVM and MXD were of 3 × 10-3 and 3.6 × 10-3 µg/L, respectively.

Silica nanoparticle-based microfluidic immunosensor with laser-induced fluorescence detection for the quantification of immunoreactive trypsin.

The purpose of this study was to develop a silica nanoparticle-based immunosensor with laser-induced fluorescence (LIF) as a detection system. The proposed device was applied to quantify the immunoreactive trypsin (IRT) in cystic fibrosis (CF) newborn screening. A new ultrasonic procedure was used to extract the IRT from blood spot samples collected on filter papers. After extraction, the IRT reacted immunologically with anti-IRT monoclonal antibodies immobilized on a microfluidic glass chip modified with 3-aminopropyl functionalized silica nanoparticles (APSN-APTES-modified glass chips). The bounded IRT was quantified by horseradish peroxidase (HRP)-conjugated anti-IRT antibody (anti-IRT-Ab) using 10-acetyl-3,7-dihydroxyphenoxazine (ADHP) as enzymatic mediator. The HRP catalyzed the oxidation of nonfluorescent ADHP to highly fluorescent resorufin, which was measured by LIF detector, using excitation lambda at 561nm and emission at 585nm. The detection limits (LODs) calculated for LIF detection and for a commercial enzyme-linked immunosorbent assay (ELISA) test kit were 0.87 and 4.2ngml(-1), respectively. The within- and between-assay variation coefficients for the LIF detection procedure were below 6.5%. The blood spot samples collected on filter papers were analyzed with the proposed method, and the results were compared with those of the reference ELISA method, demonstrating a potential usefulness for the clinical assessment of IRT during the early neonatal period.

Food safety control of zeranol through voltammetric immunosensing on Au-Pt bimetallic nanoparticle surfaces.

This study reports an accurate and sensitive strategy for zeranol (ZER) determination in bovine urine samples. ZER is a mycotoxin widely used as a synthetic growth promoter in the livestock production whose residues could present a potential risk for human health. Therefore, its use as an animal feed additive has been banned in most countries. ZER determination was accomplished using an electrochemical system in which bimetallic Au-Pt nanoparticles (Au-PtNPs) were electro-synthesized on a screen-printed carbon electrode (SPCE). The obtained Au-PtNP platform was immunofunctionalized using specific anti-ZER antibodies as a strategy to avoid potential interference. After biorecognition, ZER was directly oxidized and detected by square-wave voltammetry (SWV). The Au-PtNP surface was characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD) and cyclic voltammetry (CV). The limit of detection calculated was 0.01 ng mL(-1) with a wide linear range from 0.03 to 30 ng mL(-1). This method promises to be suitable for ZER quantification in bovine urine samples ensuring food quality and safety, as well as consumer's health.

Zinc oxide nanoparticles based microfluidic immunosensor applied in congenital hypothyroidism screening.

In this article, we present an innovative approach for congenital hypothyroidism (CHT) screening. This pathology is the most common preventable cause of mental retardation, affecting newborns around the world. Its consequences could be avoided with an early diagnosis through the thyrotropin (TSH) level measurement. To accomplish the determination of TSH, synthesized zinc oxide (ZnO) nanobeads (NBs) covered by chitosan (CH), ZnO-CH NBs, were covalently attached to the central channel of the designed microfluidic device. These beads were employed as platform for anti-TSH monoclonal antibody immobilization to specifically recognize and capture TSH in neonatal samples without any special pretreatment. Afterwards, the amount of this trapped hormone was quantified by horseradish peroxidase (HRP)-conjugated anti-TSH antibody. HRP reacted with its enzymatic substrate in a redox process, which resulted in the appearance of a current whose magnitude was directly proportional to the level of TSH in the neonatal sample. The structure and morphology of synthesized ZnO-CH NBs were characterized by scanning electron microscopy (SEM) and X-ray diffraction (XRD). The calculated detection limits for electrochemical detection and the enzyme-linked immunosorbent assay procedure were 0.00087 µUI mL(-1) and 0.015 µUI mL(-1), respectively, and the within- and between-assay coefficients of variation were below 6.31% for the proposed method. According to the cut-off value for TSH neonatal screening, a reasonably good limit of detection was achieved. These above-mentioned features make the system advantageous for routine clinical analysis adaptation.

Novel solid inks based on beeswax, graphite and graphene applied to the fabrication of paper-based sensor for galactose determination.

In this study, we present for the first time a simple and novel method for the fabrication of paper-based electrochemical sensors. The device development was carried out in a single stage with a standard wax printer. Hydrophobic zones were delimited with commercial solid ink, while electrodes were generated using new composite solid inks of graphene oxide/graphite/beeswax (GO/GRA/beeswax) and graphite/beeswax (GRA/beeswax). Subsequently, the electrodes were electrochemically activated by applying an overpotential. Various experimental variables for the GO/GRA/beeswax composite synthesis and electrochemical system obtention were evaluated. The activation process was examined by SEM, FTIR, cyclic voltammetry, electrochemical impedance spectroscopy and contact angle measurement. These studies showed morphological and chemical changes in the electrode active surface. As a result, the activation stage considerably improved the electron transfer on the electrode. The manufactured device was successfully applied for galactose (Gal) determination. This method presented a linear relation in the Gal concentration range from 84 to 1736 µmol L-1, with a LOD of 0.1 µmol L-1. The variation within and between-assay coefficients were 5.3% and 6.8%, respectively. The strategy here exposed for paper-based electrochemical sensors design is an unprecedented alternative system and represents a promising tool for mass production of economic analytical devices.

Monitoring melatonin and its isomer in Vitis vinifera cv. Malbec by UHPLC-MS/MS from grape to bottle.

Several studies have shown the presence of melatonin and related compounds in grapes and wines. The latter provides evidence of the possibility to enhance the nutraceutical properties of premium wines. However, there are many external factors that can influence the levels of this indolamine in grape and wines. In this study, the monitoring of melatonin and its tentatively identified isomer was carried out during the entire winemaking process in Vitis vinifera cv. Malbec by ultra high-performance liquid chromatography-tandem mass spectrometry. Laboratory and pilot studies were carried out to elucidate the role of grape, yeasts, and tryptophan in the evolution of the indolamines during the fermentation process. Melatonin was detected in grape extract within the range 120-160 ng/g while its isomer was found in musts and finished wines. Our results demonstrate that Saccaromyces cervisiae plays a decisive role in contributing to the content of melatonin and its isomer in wine.

A novel and rapid method for determination of natamycin in wines based on ultrahigh-performance liquid chromatography coupled to tandem mass spectrometry: validation according to the 2002/657/EC European decision.

A novel, simple, and rapid reversed-phase liquid chromatography-tandem mass spectrometric methodology was developed for the analysis of natamycin in wine samples. Natamycin was protonated to form singly charged ions in an electrospray positive ion mode. Data acquisition under MS/MS was achieved by applying multiple reaction monitoring (MRM) of three fragment ion transitions (666.3 → 648.2, 666.3 → 503.3, and 666.3 → 485.2) to provide a high degree of sensitivity and specificity. Chromatographic separation was performed on a rapid resolution column using a mobile phase consisting of an acetonitrile/water mixture with a total run time of 5.0 min. After only filtration as pretreatment, the sample was injected into the chromatographic system. The proposed method was validated in terms of selectivity, trueness, precision, decision limit (CCα), and detection capability (CCß) according to 2002/657/EC Commission decision. The values for trueness, reported as bias (%), agreed with those established by the aforementioned document. Repeatability (intraday variability) values were 12.37% at a concentration of 1.0 µg L(-1) and 8.99-4.19% at concentrations between 2.5 and 10 µg L(-1). The overall within-laboratory (interday variability) reproducibility was 15.47% at a concentration of 1.0 µg L(-1), which was significantly lower than the indicative value reported in the EU decision. The results indicated that the proposed approach is a sensitive, fast, reproducible, and robust methodology suitable for the analysis of natamycin levels in wine samples.

Development of an indirect competitive enzyme-linked immunosorbent assay applied to the Botrytis cinerea quantification in tissues of postharvest fruits.

BACKGROUND: Botrytis cinerea is a phytopathogenic fungus responsible for the disease known as gray mold, which causes substantial losses of fruits at postharvest. This fungus is present often as latent infection and an apparently healthy fruit can deteriorate suddenly due to the development of this infection. For this reason, rapid and sensitive methods are necessary for its detection and quantification. This article describes the development of an indirect competitive enzyme-linked immunosorbent assay (ELISA) for quantification of B. cinerea in apple (Red Delicious), table grape (pink Moscatel), and pear (William's) tissues. RESULTS: The method was based in the competition for the binding site of monoclonal antibodies between B. cinerea antigens present in fruit tissues and B. cinerea purified antigens immobilized by a crosslinking agent onto the surface of the microtiter plates. The method was validated considering parameters such as selectivity, linearity, precision, accuracy and sensibility. The calculated detection limit was 0.97 µg mL-1 B. cinerea antigens. The immobilized antigen was perfectly stable for at least 4 months assuring the reproducibility of the assay. The fungus was detected and quantified in any of the fruits tested when the rot was not visible yet. Results were compared with a DNA quantification method and these studies showed good correlation. CONCLUSIONS: The developed method allowed detects the presence of B. cinerea in asymptomatic fruits and provides the advantages of low cost, easy operation, and short analysis time determination for its possible application in the phytosanitary programs of the fruit industry worldwide.

Microfluidic immunosensor with gold nanoparticle platform for the determination of immunoglobulin G anti-Echinococcus granulosus antibodies.

This article describes a microfluidic immunosensor, developed for the detection of IgG antibodies specific to Echinococcus granulosus in human serum samples, which represents an alternative tool that can be used for the immunodiagnosis of hydatidosis in an automated way. Our device consists of a Plexiglas system with a central channel and a gold electrode. For immobilization of the E. granulosus antigen, a gold electrode was modified with the incorporation of gold nanoparticles. Immobilized antigen was allowed to react with IgG-anti-E. granulosus antibodies in samples, and these were quantified by horseradish peroxidase (HRP) enzyme-labeled secondary antibodies specific to human IgG using catechol (Q) as enzymatic mediator. HRP in the presence of hydrogen peroxide (H(2)O(2)) catalyzes the oxidation of Q to o-benzoquinone (P). The electrochemical reduction back to Q was detected on the gold electrode (AuE) at -0.15 V. The current obtained was proportional to the activity of the enzyme and to the concentration of antibodies of interest. The detection limit for electrochemical detection was 0.091 ng ml(-1), and the within- and between-assay coefficients of variation were below 6.7%. The proposed system presents many benefits, the more relevant are: reduced complexity and costs that are considered as the most wanted features for the clinical-immunodiagnostic field.

Determination of Ochratoxin A in apples contaminated with Aspergillus ochraceus by using a microfluidic competitive immunosensor with magnetic nanoparticles.

Ochratoxin A (OTA) is a mycotoxin produced by filamentous fungi of the genus Aspergillus and Penicillium that presents carcinogenic, teratogenic and nephrotoxic properties. In this work, we have developed, characterized and applied an immunoassay methodology comprised of magnetic nanoparticles (MNPs) as platform for immobilizing bioactive materials incorporated into a microfluidic system for rapid and sensitive quantification of Ochratoxin A (OTA) in apples (Red Delicious) contaminated with Aspergillus ochraceus. The sensor has the potential for automation and the detection of OTA was carried out using a competitive indirect immunoassay method based on the use of anti-OTA monoclonal antibodies immobilized on 3-aminopropyl-modified MNPs. The total assay time into the microfluidic competitive immunosensor was 16 min, and the calculated detection limit was 0.05 µg kg(-1). Moreover, the intra- and inter-assay coefficients of variation were below 6.5%. The proposed method can be a very promising analytical tool for the determination of OTA in apparently healthy fruits post-harvest and for its application in the agricultural industry.

A microfluidic device based on a screen-printed carbon electrode with electrodeposited gold nanoparticles for the detection of IgG anti-Trypanosoma cruzi antibodies.

In this article we report the development of an integrated microfluidic system coupled to a screen-printed carbon electrode (SPCE) applied to the quantitative determination of IgG specific antibodies present in serum samples of patients that suffer from Chagas disease. This relevant parasitic infection caused by the hemoflagellate protozoan Trypanosoma cruzi represents a major public health concern in Latin America. In order to perform the detection of mentioned antibodies, SPCE coupled to a microfluidic device was modified by electrodeposition of gold nanoparticles (AuNPs) and functionalized with Trypanosoma cruzi proteins from epimastigote membranes. The developed microfluidic immunosensor with immobilized T. cruzi proteins on the SPCE surface was successfully applied in the detection of specific IgG anti-T. cruzi antibodies, which were allowed to react immunologically with immobilized T. cruzi antigen. After that, labelled antibodies were quantified through the addition of horseradish peroxidase (HRP) enzyme-labeled secondary antibodies specific to human IgG, using 4-tert-butylcatechol (4-TBC) as enzymatic mediator. HRP in the presence of hydrogen peroxide (H(2)O(2)) catalyzes the oxidation of 4-TBC whose back electrochemical reduction was detected on a modified electrode at -100 mV. The calculated detection limit for electrochemical detection was 3.065 ng mL(-1) and the intra- and inter-assay coefficients of variation were below 6.95%.

Online immunoaffinity assay-CE using magnetic nanobeads for the determination of anti-Helicobacter pylori IgG in human serum.

About two-thirds of the world's population is infected with Helicobacter pylori (H. pylori). This Gram-negative bacterium is the most important etiological agent of chronic active type B gastritis and peptic ulcer diseases. Conventional methods such as gastric biopsy, ELISA and culture, require a long time for the determination of H. pylori infections. Moreover, the antibodies in human serum sample are capable to react immunologically with the purified H. pylori antigens immobilized on different kinds of support like magnetic nanobeads. In this study, we have developed an online immunoaffinity assay-CE to determine the concentration of anti-H. pylori IgG using magnetic nanobeads as a support of the immunological affinity ligands and an LIF as a detector. The separation was performed in 0.1 M glycine-HCl, pH 2, as the background electrolyte. The linear calibration curve to predict the concentration of H. pylori-specific immunoglobulin G antibodies in serum was produced within the range of 0.12-100 U/mL. The linear regression equation was i = 492.86+96.03 × C(anti-H. pylori), with the linear regression coefficient r(2) = 0.999. The LOD calculated by fluorescence detection procedure was of 0.06 U/mL. The whole assay was done in no more than 35 min and it was entirely automatized. The development of immunoaffinity assay-CE in this study demonstrates that there is a large possibility to introduce nanotechnology in several fields with significant advantages over the classic methodologies. Our proposition comprises the diagnosis and screening field.

IgG anti-gliadin determination with an immunological microfluidic system applied to the automated diagnostic of the celiac disease.

In the present article, a novel microfluidic immunosensor coupled with electrochemical detection for anti-gliadin IgG antibody quantification is proposed. This device represents an important tool for a fast, simple, sensitive, and automated diagnostic for celiac disease, which is carried out through detection of anti-gliadin IgG antibodies present in human serum samples. Celiac disease (CD) is an autoimmune disease generated by gluten protein fractions called prolamins. This pathology affects about one in 250 people around the world, produces intestinal inflammation, villous atrophy, and crypt hyperplasia, which causes a range of symptoms including altered bowel habits, malnutrition and weight loss. Our immunosensor consists of a Plexiglas device coupled to a gold electrode, with a central channel containing 3-aminopropyl-modified controlled pore glass (AP-CPG). The quantification of anti-gliadin IgG antibodies was carried out using a heterogeneous, non-competitive enzyme-linked immunosorbent assay (ELISA) in which IgG antibodies bound to gliadin protein, immobilized on AP-CPG, were determined by alkaline phosphatase (AP) enzyme-labeled second antibodies specific to human IgG. The p-aminophenyl phosphate (p-APP) was converted to p-aminophenol (p-AP) by AP, and the electroactive product was quantified on a gold electrode at 0.250 V. The calculated detection limits for electrochemical detection and the ELISA procedure were 0.52 and 2.72 UR mL(-1), respectively, and the within- and between-assay coefficients of variation were below 5.8%. The optimized procedure was applied to the determination of anti-gliadin IgG antibodies in human serum samples.

Determination of beta-glucosidase activity in soils with a bioanalytical sensor modified with multiwalled carbon nanotubes.

Soil microorganisms and enzymes are the primary mediators of soil biological processes, including organic matter degradation, mineralization, and nutrient recycling. They play an important role in maintaining soil ecosystem quality and functional diversity. Moreover, enzyme activities can provide an indication of quantitative changes in soil organic matter. Beta-glucosidase (beta-Glu) activity has been found to be sensitive to soil management and has been proposed as a soil quality indicator because it provides an early indication of changes in organic matter status and its turnover. The aims of the present study were to test and use a simple and convenient procedure for the assay of beta-Glu activity in agricultural soil. The method described here is based on the enzymatic degradation of cellobiose by beta-Glu present in the soil sample and the subsequent determination of glucose produced by the enzymatic reaction using screen-printed carbon electrodes modified with multiwalled carbon nanotubes (SPCE-CNT) equipped with coimmobilized glucose oxidase and horseradish peroxidase enzymes. The potential applied to the SPCE-CNT detection was -0.15 V versus a Ag/AgCl pseudo-reference electrode. A linear calibration curve was obtained in the range 2.7-11.3 mM with a correlation coefficient. In the present study, an easy and effective SPCE-CNT-modified electrode allowed an improved amperometric response to be achieved and this is attributed to the increased surface area upon electrode modification.

Electrochemical microfluidic immunosensor based on TES-AuNPs@Fe3O4 and CMK-8 for IgG anti-Toxocara canis determination.

We report a microfluidic immunosensor for the electrochemical determination of IgG antibodies anti-Toxocara canis (IgG anti-T. canis). In order to improve the selectivity and sensitivity of the sensor, core-shell gold-ferric oxide nanoparticles (AuNPs@Fe3O4), and ordered mesoporous carbon (CMK-8) in chitosan (CH) were used. IgG anti-T. canis antibodies detection was carried out using a non-competitive immunoassay, in which excretory secretory antigens from T. canis second-stage larvae (TES) were covalently immobilized on AuNPs@Fe3O4. CMK-8-CH and AuNPs@Fe3O4 were characterized by transmission electron microscopy, scanning electron microscopy, energy dispersive spectrometry, cyclic voltammetry, electrochemical impedance spectroscopy, and N2 adsorption-desorption isotherms. Antibodies present in serum samples immunologically reacted with TES, and then were quantified by using a second antibody labeled with horseradish peroxidase (HRP-anti-IgG). HRP catalyzes the reduction from H2O2 to H2O with the subsequent oxidation of catechol (H2Q) to p-benzoquinone (Q). The enzymatic product was detected electrochemically at _100 mV on a modified sputtered gold electrode. The detection limit was 0.10 ng mL-1, and the coefficients of intra- and inter-assay variation were less than 6%, with a total assay time of 20 min. As can be seen, the electrochemical immunosensor is a useful tool for in situ IgG antibodies anti-T. canis determination.