Effects of redox conditions and zinc(II) ions on metallothionein aggregation revealed by chip capillary electrophoresis

Metallothioneins (MTs) belong to cysteine-rich proteins with unique higher structure. One of the most known MT's functions is metals detoxification and maintaining their homeostasis in a cell. Structure of MT with naturally occurred zinc(II) ions can be affected by concentration of metal ions as well as redox milieu inside a cell, however the exact explanation and biochemical effects of the structural changes are still missing. In this study we used capillary electrophoresis on chip coupled with fluorescence detection to determine structural changes of MT with increasing concentration of zinc(II) ions and under various redox conditions. To investigate the structural-dependent effects, reduced and/or oxidized apo-MT (MT without natural occurred metal ion) was prepared. Zinc binding into reduced and/or oxidized apo-MT was compared. MT was incubated with 0, 5, 15, 25, 50 and 100 μM ZnCl₂ for 1 h in 37 °C. Formation of MT aggregates with increasing zinc concentration was observed by spectrophotometry, chip capillary electrophoresis, and SDS-PAGE. We found out that reduced MT forms aggregates more readily compared to oxidized MT. Using the chip capillary electrophoresis allowed us relative quantification of MT aggregation as a decrease in the area of the signal corresponding to the monomer form of MT (Mw 15 kDa, migration time 26.5 s) and its ratio to total signal (sum of all signals measured by the electrophoresis). The dependences had an exponential character with equation y = 2.4 × e^−0.01x, R² = 0.945 for 15 kDa peak area and y = 0.11 × e^−0.01x, R² = 0.938 for decrease of 15 kDa peak area ratio to the total signal. Zn–MT interaction was 30% faster during the first 15 min and 50% faster during the whole experiment for reduced MT. It can be concluded that formation of MT aggregates is dependent on redox state and Zn(II) concentration.     

Chip electrospray mass spectrometry for carbohydrate analysis

Currently two types of chip systems are used in conjunction with MS: out-of-plane devices, where hundreds of nozzles, nanospray emitters are integrated onto a single silicon substrate from which electrospray is established perpendicular to the substrate, and planar microchips, embedding a microchannel at the end of which electrospray is generated in-plane, on the edge of the microchip. In the last two years, carbohydrate research greatly benefited from the introduction and implementation of the chip-based MS. In two laboratories the advantages of the chip electrospray in terms of ionization efficiency, sensitivity, reproducibility, quality of data in combination with high mass accuracy, and resolution of detection were systematically explored for several carbohydrate classes: O- and N-glycopeptides, oligosaccharides, gangliosides and glycoprotein-derived O- and N-glycans, and glycopeptides. The current state-of-the-art in interfacing the chip electrospray devices to high-performance MS for carbohydrate analysis, and the particular requirements for method optimization in both positive and negative ion modes are reviewed here. The recent applications of these miniaturized devices and their general potential for glycomic-based surveys are highlighted.     

A new micro-fluid chip calorimeter for biochemical applications

Based on a new thermopile heat power sensor, which was developed in MEMS technology, a miniaturized flow-through calorimeter was constructed. The heat power sensor consists of a silicon chip with a thin film BiSb/Sb thermopile and a PMMA reaction chamber. To ensure high signal resolution the heat power sensor is mounted inside a high-precision thermostat, which has a temperature stability of less than 100 μK. The heat power sensitivity of the calorimeter is 4-7 V W⁻¹ depending on the thermal conductivity of the liquid, the height of the chosen reaction chamber and the volume flow rate. A limit of detection of less than 50 nW can be obtained for volume flows lower than 5 μl min⁻¹. An important advantage is the low sample consumption of the calorimeter. For special applications the sample need for one measurement pulse does not exceed 20 μl.     

DNA修饰电极的研究(Ⅸ)--DNA探针在金基底上的固定、表征及其表面分子杂交

采用疏基化合的自组装／共价键合反应的逐层固定方法将双链DNA固定到金表面得到DNA修饰电极,并对该电极表面进行了电化学和X射线光电子能谱表征。研究了电极表面固定化DNA的表面分子杂交。对开发电化学基因诊断芯片和基因传感器具有一定意义。     

Photobleaching-based flow measurement in a commercial capillary electrophoresis chip instrument

For microfluidic analytical instruments, a facile, fast, and accurate instrument test is highly demanded. The test includes the quantitative verification of the relationship between pressure drop and flow velocity for the hydrodynamic pump, between the electric voltage and electroosmotic flow (EOF) for the high-voltage supply, and the chip quality. The key point for the test is the measurement of the flow velocity. However, most currently available velocimetries cannot be directly used without any instrumental modification or adding extra instruments. We applied a recently developed Laser Induced Fluorescence Photobleaching Anemometer (LIFPA) for the instrument test through measuring fluid flow velocity in a microfluidic instrument with optical measurement without any modification and extra instrument. We have successfully used the method to test Caliper HTS 250 System from Caliper Life Sciences (Hopkinton, MA) with its own light source and detector. The experimental result demonstrates that this single-point method of measuring flow velocity can be easily used for accurate test of a microfluidic instrument in less than 10 min at extremely low cost without any modification and extra instrument.     

Surface acoustic wave biosensors: a review

This review presents an overview of 20 years of worldwide development in the field of biosensors based on special types of surface acoustic wave (SAW) devices that permit the highly sensitive detection of biorelevant molecules in liquid media (such as water or aqueous buffer solutions). 1987 saw the first approaches, which used either horizontally polarized shear waves (HPSW) in a delay line configuration on lithium tantalate (LiTaO₃) substrates or SAW resonator structures on quartz or LiTaO₃ with periodic mass gratings. The latter are termed “surface transverse waves” (STW), and they have comparatively low attenuation values when operated in liquids. Later Love wave devices were developed, which used a film resonance effect to significantly reduce attenuation. All of these sensor approaches were accompanied by the development of appropriate sensing films. First attempts used simple layers of adsorbed antibodies. Later approaches used various types of covalently bound layers, for example those utilizing intermediate hydrogel layers. Recent approaches involve SAW biosensor devices inserted into compact systems with integrated fluidics for sample handling. To achieve this, the SAW biosensors can be embedded into micromachined polymer housings. Combining these two features will extend the system to create versatile biosensor arrays for generic lab use or for diagnostic purposes. SAW based biosensor immersed in a sample flow. Analyte molecules binding to the immobilized antibodies on the sensor surface will influence the velocity of the SAW and hence the output signal generated by the driving electronics.     

不同沉积层的纳米波导谐振腔特性测试

为确定硅基片上系统半导体光电器件集成中绝缘层材料对器件整体性能的影响,设计并制备了带有覆层的纳米波导谐振腔.谐振透射谱功率测试表明顶层覆盖Si_3N_4薄膜和SiO_2薄膜绝缘层没有削弱环形谐振腔的品质因素,沉积后的最佳耦合间距为70~110 nm.覆层为SiO_2时谐振点波长附近的谐振峰消光比达16.5 dB,3 dB带宽为0.12 nm;覆层为Si_3N_4时谐振点波长附近的谐振峰消光比达13.9 dB,3 dB带宽为0.18 nm.该研究为片上系统集成设计中最佳绝缘层材料的选择提供参考.     

Lamination-based rapid prototyping of microfluidic devices using flexible thermoplastic substrates

Transposing highly sensitive DNA separation methods (such as DNA sequencing with high read length or the detection of point mutations) to microchip format without loss of resolution requires fabrication of relatively long (approx. 10 cm) microchannels along with sharp injection bands. Conventional soft lithography methods, such as mold casting or hot-embossing in a press, are not convenient for fabricating long channels. We have developed a lamination-based replication technique for rapid fabrication of sealed microfluidic devices with a 10 cm long, linear separation channel. These devices are fabricated in thin cyclo-olefin copolymer (COC) plastic substrates, thus making the device flexible and capable of assuming a range of 3-D configurations. Due to the good optical properties of COC, this new family of devices combines multiple advantages of planar microfluidics and fused-silica capillaries.     

The potential of luminescence signals from electronic components for accident dosimetry

This study investigated the optically stimulated luminescence of a large number of electronic components extracted from both old and new generation mobile phones and chip modules of phone cards. Most resistors and all chip modules studied present a linear dose response (R > 0.99) in the dose range investigated (200 mGy up to 6 Gy, respectively 10 Gy), while capacitors, inductors and integrated circuits generally have a non-linear growth (exponential or cubic). For our experimental setup, an average specific luminescence of ∼20,000 cts in 2 s/Gy (n = 10) and ∼6000 cts in 2 s/Gy (n = 14) was obtained for two types of chip modules with a relatively high degree of homogeneity (relative standard deviation of 23% and 31%) and a minimum detectable dose of 7 mGy for immediate measurement. The investigated signals show small sensitivity changes (generally <10%) after repeated cycles of irradiation and readout. Preliminary fading measurements are presented. It can be concluded that most mobile phones and phone card components have a significant potential as retrospective luminescence dosimeters.