Effect of surface roughness on performance of magnetoelastic biosensors for the detection of Escherichia coli.

Escherichia coli are bacteria that must be controlled in the food industry and the hospital sector. Magnetoelastic biosensors offer the promise of rapid identification of these and other harmful antigens. In this work, strips of amorphous Metglas 2826MB3 were cut to size (5 mm × 1 mm) with a microdicing saw and were then coated with thin layers of Cr and Au, as verified by Rutherford backscattering spectroscopy (RBS). Several sensor surfaces were studied: 1) as-cast strip, wheel side; 2) as-cast strip, free surface; and 3) thinned and polished surface. A layer of cystamine was applied to the Au-covered magnetoelastic substrate, forming a self-assembledmonolayer (SAM), followed by antibodies, using a modified Hermanson protocol. The cystamine layer growth was verified by Fourier transform infrared spectroscopy (FTIR) and scanning electronmicroscopy (SEM). The biosensors were exposed to solutions of bacteria and the resonant frequency of the sensors was measured with an impedance analyzer for times up to 100 min. Reductions in the resonant frequency, corresponding to bacteria capture, were measured after optimizing the signal amplitude. For times up to 40 min, high capture rates were observed and thereafter saturation occurred. Saturation values of the frequency shifts were compared with the number of bacteria observed on the sensor using fluorescence microscopy. Parameters associated with capture kinetics were studied for different sensor surfaces. The rough surfaces were found to show a faster response, while the thinned and polished sensors showed the largest frequency shift.

A nanostructured piezoelectric immunosensor for detection of human cardiac troponin T.

A piezoelectric immunosensor based on gold nanoparticles (AuNPs) co-immobilized on a dithiol-modified surface is proposed for detection of human cardiac troponin T (TnT). Anti-human troponin T (anti-TnT) antibodies were covalently immobilized on the nanostructured electrode surface by thiol-aldehyde linkages. In a homogeneous bulk solution, TnT was captured by anti-TnT immobilized on the QCM electrode. Cyclic voltammetry studies were used to characterize the AuNPs layer on the electrode surface and the anti-TnT immobilization steps. The QCM-flow immunosensor exhibited good reliability, measuring concentrations of TnT from 0.003 to 0.5 ng mL(-1) in human serum with high linearity (r = 0.989; p < 0.01). The immunosensor exhibited a 7% coefficient of variation and 0.0015 ng mL(-1) limit of detection, indicating a high reproducibility and sensitivity. The proposed QCM nanostructured immunosensor is easy to use and has promising potential in the diagnosis of acute myocardial infarction due to its speed and high sensitivity.

Methylenetetrahydrofolate reductase gene polymorphism and its association with coronary artery disease.

CONTEXT AND OBJECTIVE: Obstructive coronary artery disease (CAD) is characterized by the deposition of atherosclerotic plaque on the coronary artery wall. Its manifestations depend on interactions between environmental and genetic risk factors. The aim of this work was to analyze the frequency of methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism in patients with CAD and its association with plasma homocysteine levels. Risk factors for CAD were also evaluated. DESIGN AND SETTING: Retrospective with blind quantitative analysis, at Hospital de Base, Faculdade de Medicina de São José do Rio Preto. METHODS: One hundred and twenty-seven individuals were studied. All completed a questionnaire to analyze risk factors for CAD. MTHFR polymorphism was investigated by restriction fragment length analysis and correlated with the number of affected arteries and degree of arterial obstruction determined by coronary cineangiography, and with plasma homocysteine levels measured by liquid chromatography/sequential mass spectrometry. RESULTS: Smoking (p = 0.02) and high-density lipoprotein cholesterol (p = 0.01) were associated with CAD. The C allele was the most prevalent in patients (0.61) and controls (0.66). There was no correlation between MTHFR/C677T polymorphism and plasma homocysteine levels. However, in patients with the TT genotype there was a correlation with the prevalence of coronary obstruction greater than 95% (p = 0.02) and the presence of two affected arteries (p = 0.04). CONCLUSIONS: The TT genotype is associated with coronary artery obstruction greater than 95% and the presence of two affected arteries. This confirms the relationship between genetic variants in specific patient subgroups and cardiovascular diseases.

Methylenetetrahydrofolate reductase gene polymorphism and its association with coronary artery disease

CONTEXT AND OBJECTIVE: Obstructive coronary artery disease (CAD) is characterized by the deposition of atherosclerotic plaque on the coronary artery wall. Its manifestations depend on interactions between environmental and genetic risk factors. The aim of this work was to analyze the frequency of methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism in patients with CAD and its association with plasma homocysteine levels. Risk factors for CAD were also evaluated. DESIGN AND SETTING: Retrospective with blind quantitative analysis, at Hospital de Base, Faculdade de Medicina de São José do Rio Preto. METHODS: One hundred and twenty-seven individuals were studied. All completed a questionnaire to analyze risk factors for CAD. MTHFR polymorphism was investigated by restriction fragment length analysis and correlated with the number of affected arteries and degree of arterial obstruction determined by coronary cineangiography, and with plasma homocysteine levels measured by liquid chromatography/sequential mass spectrometry. RESULTS: Smoking (p = 0.02) and high-density lipoprotein cholesterol (p = 0.01) were associated with CAD. The C allele was the most prevalent in patients (0.61) and controls (0.66). There was no correlation between MTHFR/C677T polymorphism and plasma homocysteine levels. However, in patients with the TT genotype there was a correlation with the prevalence of coronary obstruction greater than 95 percent (p = 0.02) and the presence of two affected arteries (p = 0.04). CONCLUSIONS: The TT genotype is associated with coronary artery obstruction greater than 95 percent and the presence of two affected arteries. This confirms the relationship between genetic variants in specific patient subgroups and cardiovascular diseases.

CONTEXTO E OBJETIVO: A doença arterial coronariana (DAC) caracteriza-se pelo depósito de placa aterosclerótica na parede arterial coronária. Sua manifestação é dependente da interação entre fatores de risco ambientais e genéticos. O objetivo deste trabalho é analisar a freqüência do polimorfismo MTHFR/C677T em pacientes com doença arterial coronária e sua associação com o nível de Hcy plasmática. Fatores de risco para DAC também foram avaliados. TIPO DE ESTUDO: Retrospectivo com análise cega quantitativa, no Hospital de Base, Faculdade de Medicina de São José do Rio Preto. MÉTODOS: Foram estudados 127 indivíduos. Todos responderam a um questionário para análise dos fatores de risco para DAC. O polimorfismo MTHFR/C677T, investigado por análise de comprimento de fragmentos de restrição, foi correlacionado com número de artérias afetadas e grau de obstrução arterial, determinadas pela cinangiocoronariografia, e com o nível de Hcy plasmática. RESULTADOS: Tabagismo (p = 0,02) and HDLc (p = 0,01) foram associados com DAC. O alelo C foi o mais prevalente em pacientes (0,61) e controles (0,66; p = 0,49). O polimorfismo MTHFR/C677T não apresentou associação com níveis de Hcy plasmática. Entretanto, nos pacientes com genótipo TT observou-se a prevalência de obstrução coronariana acima de 95 por cento (p = 0,02) e a presença de duas artérias lesadas (p = 0,04). CONCLUSÕES: Associou-se o genótipo TT com o grau de obstrução arterial coronária acima de 95 por cento e a presença de duas artérias lesadas; confirma-se, assim, a relação de variantes genéticas em subgrupos específicos de pacientes com doenças cardiovasculares.

Involvement of transglutaminase in the formation of covalent cross-links in the cell wall of Candida albicans.

Activity of the enzyme glutaminyl-peptide--glutamylyl-transferase (EC 2.3.2.13; transglutaminase), which forms the interpeptidic cross-link N epsilon-(gamma-glutamic)-lysine, was demonstrated in cell-free extracts obtained from both the yeast like and mycelial forms of Candida albicans. Higher levels of enzymatic activity were observed in the cell wall fraction, whereas the cytosol contained only trace amounts of activity. Cystamine, a highly specific inhibitor of the enzyme, was used to analyze a possible role of transglutaminase in the organization of the cell wall structure of the fungus. Cystamine delayed protoplast regeneration and inhibited the yeast-to-mycelium transition and the incorporation of proteins into the cell wall. The incorporation of covalently bound high-molecular-weight proteins into the wall was sensitive to cystamine. Proteic epitopes recognized by two monoclonal antibodies, one of which is specific for the mycelial walls of the fungus, were also sensitive to cystamine. These data suggest that transglutaminase may be involved in the formation of covalent bonds between different cell wall proteins during the final assembly of the mature cell wall.

Rat duodenum nitrergic-induced relaxations are cGMP-independent and apamin-sensitive.

The effects of the K+ channel blockers, apamin, tetraethylammonium and 4-aminopyridine, upon the relaxations of the isolated rat proximal duodenum induced by nitregic nerve activation, nitric oxide (NO), the NO donor 3-morpholinosydnonimine (SIN-1) and Br-cyclic GMP were determined. The effects of the guanylate cyclase inhibitors, cystamine and N-methylhydroxylamine, on NO-, SIN-1- and nitrergic nerve-induced responses were also investigated. Apamin inhibited nitrergic nerve-, NO-and SIN-1-induced relaxations but did not affect those induced by Br-cGMP. Tetraethylammonium and 4-aminopyridine as well as cystamine and N-methylhydroxylamine failed to affect the relaxations caused by any of the agents tested. These findings indicate that, in the rat proximal duodenum, nitrergic nerve activation as well as exogenous nitric oxide cause relaxation through a cGMP-independent, apamin sensitive mechanism.