生物传感器与生物电子学

Background: Recent studies suggest that elevated dopamine concentrations reduce mitochondrial membrane potential, potentially contributing to neurological or other disorders. Changes in average membrane potential of suspended cells or organelles can be monitored in real time by measuring impedance of the suspension as a labelfree assay. Methods/Results: Here we report on measurements of frequency- and time-dependent electrical impedance of mitochondrial suspensions in the presence of various substrates, including dopamine. We find that, during succinateinduced state-2 respiration, addition of either trifluoromethoxy carbonylcyanide phenylhydrazone (FCCP), a known mitochondrial uncoupler (depolarizer), or dopamine at high concentrations cause similar increases in impedance, consistent with a drop in mitochondrial membrane potential. Conclusion: Our results provide further evidence that dopamine reduces mitochondrial membrane potential, albeit less severely than FCCP, possibly by generating free radicals that partially perforate the mitochondrial inner membrane and/or reduce activity of respiratory complexes. These results suggest that prolonged excess dopamine can impair ATP production, with possible implications in some neurological disorders and suggesting the need for care in the use of dopamine to treat Parkinson’s disease or heart failure.

We present the development and simplification of label-free fiber optic biosensors based on immobilization of oligonucleotides on dual-peak long period gratings (dLPGs). This improvement is the result of a simplification of biofunctionalization methodology. A one-step 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)-mediated reaction has been developed for the straightforward immobilization of unmodified oligonucleotides on the glass fiber surface along the grating region, leading to covalent attachment of a 5´-phosphorylated probe oligonucleotide to the amino-derivatized fiber grating surface. Immobilization is achieved via a 5´phosphate-specific linkage, leaving the remainder of the oligonucleotide accessible for binding reactions. The dLPG has been tested in different external media to demonstrate its inherent ultrahigh sensitivity to the surrounding-medium refractive index (RI) achieving 50- fold improvement in RI sensitivity over the previously-published LPG sensor in media with RI’s relevant to biological assays. After functionalization, the dLPG biosensor was used to monitor the hybridization of complementary oligonucleotides showing a detectable oligonucleotide concentration of 4 nM. The proposed one-step EDC reaction approach can be further extended to develop fiber optic biosensors for disease analysis and medical diagnosis with the advances of label-free, real-time, multiplex, high sensitivity and specificity.

A sensor chip for specific detection of blood glucose was developed. The sensor utilized a surface-plasmonresonance (SPR) setup on a prism based Kretschmann configuration utilizing spectral interrogation scheme. Selfassembled monolayers (SAMs) based preparation provided the chip stability due to covalent bonds and hence it can be reused for multiple times. Such a scheme can be advantageous in continuous monitoring of blood glucose, without changing the chip. Control experiments were performed without molecular recognition layer to confirm the performance of the sensor. Furthermore, the measurements were performed on blood serums and compared with that of a conventional glucometer. The present sensor has the advantages of improved sensitivity (0.14 nm/(mg/dl)) and stable response for 3 months, which are better compared to existing reports.

Development of a rapid and label-free Electrochemical Impedance Spectroscopy (EIS) biosensor for nucleic acid detection based on sensitivity and specifies was proposed to fill the technology gap between rapid and portable detection of DNA sequences. Current biosensors have inherent limitations to sensitivity and specificity due to the monovalent character of the detection method. These limitations were significantly improved upon by constructing cooperative probes, called Tentacle Probes™ (TP), which utilize divalent binding to achieve both high sensitivity as well as extremely high specificity. Immobilization of Tentacle Probe onto gold disk electrode produced statistical significance, with p<3.0*10-13. Tentacle Probe with EIS detection was able to detect a difference between the B. AnthracisgyrA gene and B. Cereus gyrA Single Nucleotide Polymorphism sequence at a lower limit of detection of 20nM. Furthermore, B. Cereus gyrA is a common soil bacterium that causes false positives for B. AnthracisgyrA. This method of detection is current a model system for DNA detection; other genes could be detected with this system. With the potential to be transformative by providing a platform technology for both DNA and protein detection with high sensitivity this system offers extremely low rates of false positives. There is a high capacity for the technology to be multiplexed for high-throughput applications or quick diagnostic tests for medical purposes. Thus, the biomarker development will also have the potential to be assessed through noninvasive methods.

In this paper, a three-layer artificial neural network (ANN) was investigated to predict the inhibitory concentration (IC) values assessed via MTT cell viability assay on the four types of human lung epithelial cancer cell lines. In order to achieve this purpose, a multilayer perceptron (MLP) neural network trained with back-propagation algorithm was employed for developing the ANN model. To develop the model, the input parameters were concentrations and types of cell lines and the outputs were IC10, IC20, IC30, IC40, IC50, IC60, IC70 and IC80 values in the A549, H157, H460 and H1975 cell lines. The proposed ANN model has achieved good agreement with the experimental data and has a small error between the estimated and experimental values. The obtained results show that the proposed ANN model is a useful, reliable, fast and cheap tool to predict the IC values assessed via MTT cell viability assays.

The special attention is given to methods for control of genotoxicity. Among them, the detailed analysis is given that approaches which are based on the traditional molecular genetics tests, common instrumental systems and a modern biosensor devices. It is provided as general characteristics of the developed widely dispersed, most applicable in the practice approaches and detailed description of the basic principles of their functioning. The last a specially concerns instrumental analytical methods and, in particular, cell biosensors considering the possible type of the transducers, types of cells as sensitive structures, their integration in sensor elements and way of specific signal registration. In general it is analysed sensitivity and field of application of the existed approaches for the control of total toxicity and genotoxicity.

An electrochemical simultaneous evaluation of dopamine (DA), serotonin (5-HT) and ascorbic acid (AA) in phosphate buffer solution (PBS) of pH 7.4 was attempted using a cationic surfactant, tetraoctylammonium bromide, modified carbon paste electrode (TOABMCPE). At the modified electrode, a well-defined redox peak with a great enhancement in current was witnessed. The oxidation of DA was found to be greatly pH dependent. The TOAB fabricated electrode overcomes the problem of resolution of DA with the coexisting species, AA and 5-HT. Simultaneous studies through cyclic voltammetric and differential voltammetric techniques gave excellent results with a great potential difference between DA-AA and DA-5-HT. The detection limit of the modified electrode was found to be 0.019 μM with the aid of amperometry. The developed sensor was applied for the detection of DA in injection samples.

In this paper, we report an original method to immobilize gold nanorods onto mixed self-assembled monolayers (SAMs) of Mercaptoundecanoic Acid (MUA), Mercaptohexanol (MOH) and cysteamine (CYS) onto planar gold surface. Polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS), scanning electron microscopy (SEM) and transmission electron microscopy (TEM) images revealed a remarkable shape and size narrow distribution on well functionalized gold surfaces, as well as a tendency to form linear assemblies after immobilization. The results highlight the good distribution of gold nanorods with an average length of 32.6 ± 0.9 nm and width of 13 ± 1.8 nm, the simplicity of the immobilization procedure of gold nanorods and the interest of using them as labels to enhance the sensitivity of FT-SPR-based sensors. gold nanostructured surface FT-SPR measurements of biorecognition using gold nanorods immobilized onto gold surfaces were performed at various prostatic antigen specific (PSA) concentrations, from 5 mg/L to 0.5 mg/L reaching a system sensitivity of 37 ± 2 cm-1/ mgL-1.

Milk is a bulk consumer product therefore it serves a versatile source of public exposure to contaminants. Among the various heavy metals, lead has been recognized as the leading environmental health threat and milk has been found mostly contaminated with high levels of Pb (II) ions. In present study an optical biosensor is developed employing Bacillus sphaericus (MTCC 5100) as biorecognition unit and the analysis is based on urease inhibition resulting in fluorescence change. Investigation was carried out by means of fluorescence dye (Rhodamine 6G) in hydrophobic environment with the heavy metal. The novelty of the method lies in the formula that is devised to detect equivalent Pb (II) ions in milk in the presence of Cd (II). Lowest detection limit achieved is 0.48 nM Pb (II) equivalents in spiked milk samples (permissible limit 96.6 nM). This is the first report on low level Pb (II) monitoring in milk through high throughput microarray biosensing.

In the present study, highly monodispersed polyaniline (PANI) and gold nanoparticles grafted PANI nanospheres (Au@PANI) have been fabricated through a facile wet-chemical synthesis process by in-situ polymerization of aniline. The structural characterization of the PANI and Au@PANI nanospheres were carried out by UV-Vis spectroscopy, FT-IR spectroscopy, and High Resolution Transmission Electron Microscopy (HR-TEM). Stable uniform sized nanospheres of PANI and Au@PANI obtained were found to exhibit superior gas sensor performance towards trace-level concentration of ammonia (NH3) gas in the range of 1-5 ppm. A comparative analysis of the sensor properties of PANI and Au@PANI nanospheres were performed under ambient conditions. Au@PANI nanospheres exhibited enhanced gas sensing performance with high sensitivity (58.2%), rapid response (48 s) and recovery (58 s) characteristics (1 ppm of NH3). The results indicate the potential use of Au@PANI nanospheres for fabrication of efficient NH3 sensors for varied applications.