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Yakugaku Zasshi : Journal of the... Nov 2008Psychological stress is of major importance to all age groups in recent years, and may lead to mental disorder and various diseases. An objective and quantitative method...
Psychological stress is of major importance to all age groups in recent years, and may lead to mental disorder and various diseases. An objective and quantitative method for measuring salivary stress-related substances is highly desired because saliva collection is easy, stress free and noninvasive. We have developed a rapid and easy-to-use analytical tool for the measurement of cortisol and secretory immunoglobulin A (sIgA) based on microchip technology, immunoselectivity and electrophoretic separation technique. Performing immunoreaction and capillary electrophoresis (CE) separation on microchips is a promising technique for on-site determination of biogenic substances, and has a few advantages over conventional immunoassay methods: reduced sample size, shortening analysis times, high separation efficiency, reduced cost, and downsizing of analytical system. At this stage of our research, some preliminary prototypes of a high-sensitive microchip CE instrument were constructed to determine the stress-related substances in real saliva samples. However, there is not enough detection sensitivity for cortisol analysis. On the other hand, sIgA was successfully analyzed using a laboratory-built microchip CE system and optimal analytical conditions. The sIgA determination is rapid compared with a conventional immunoassay method, and provides an acceptable degree of repeatability and recovery. In the future, microchip technologies will enable total automation and integration of sample preparation. This research has widespread future potential for monitoring multiple stress-related markers within minutes from a trace of saliva, and can contribute to disease prevention and overall good health.
Topics: Biomarkers; Electrophoresis, Microchip; Humans; Hydrocortisone; Immunoglobulin A, Secretory; Reproducibility of Results; Saliva; Stress, Psychological
PubMed: 18981695
DOI: 10.1248/yakushi.128.1595 -
Journal of Biomedicine & Biotechnology 2008A compact integrated system-on-chip (SoC) architecture solution for robust, real-time, and on-site genetic analysis has been proposed. This microsystem solution is...
A compact integrated system-on-chip (SoC) architecture solution for robust, real-time, and on-site genetic analysis has been proposed. This microsystem solution is noise-tolerable and suitable for analyzing the weak fluorescence patterns from a PCR prepared dual-labeled DNA microchip assay. In the architecture, a preceding VLSI differential logarithm microchip is designed for effectively computing the logarithm of the normalized input fluorescence signals. A posterior VLSI artificial neural network (ANN) processor chip is used for analyzing the processed signals from the differential logarithm stage. A single-channel logarithmic circuit was fabricated and characterized. A prototype ANN chip with unsupervised winner-take-all (WTA) function was designed, fabricated, and tested. An ANN learning algorithm using a novel sigmoid-logarithmic transfer function based on the supervised backpropagation (BP) algorithm is proposed for robustly recognizing low-intensity patterns. Our results show that the trained new ANN can recognize low-fluorescence patterns better than an ANN using the conventional sigmoid function.
Topics: Biomimetics; Electronics; Equipment Failure; Equipment Failure Analysis; Gene Expression Profiling; Pattern Recognition, Automated; Polymerase Chain Reaction; Signal Processing, Computer-Assisted; Spectrometry, Fluorescence
PubMed: 18566679
DOI: 10.1155/2008/259174 -
Proceedings of the National Academy of... Apr 2011The metastatic invasion of cancer cells from primary tumors to distant ecological niches, rather than the primary tumors, is the cause of much cancer mortality [Zhang...
The metastatic invasion of cancer cells from primary tumors to distant ecological niches, rather than the primary tumors, is the cause of much cancer mortality [Zhang QB, et al. (2010) Int J Cancer 126:2534-2541; Chambers AF, Goss PE (2008) Breast Cancer Res 10:114]. Metastasis is a three-dimensional invasion process where cells spread from their site of origin and colonize distant microenvironmental niches. It is critical to be able to assess quantitatively the metastatic potential of cancer cells [Harma V, et al. (2010) PLoS ONE 5:e10431]. We have constructed a microfabricated chip with a three-dimensional topology consisting of lowlands and isolated square highlands (Tepuis), which stand hundreds of microns above the lowlands, in order to assess cancer cell metastatic potential as they invade the highlands. As a test case, the invasive ascents of the Tepui by highly metastatic PC-3 and noninvasive LNCaP prostate cancer cells were used. The vertical ascent by prostate cancer cells from the lowlands to the tops of the Tepui was imaged using confocal microscopy and used as a measure of the relative invasiveness. The less-metastatic cells (LNCaP) never populated all available tops, leaving about 15% of them unoccupied, whereas the more metastatic PC-3 cells occupied all available Tepuis. We argue that this distinct difference in invasiveness is due to contact inhibition.
Topics: Cell Line, Tumor; Humans; Male; Microchip Analytical Procedures; Neoplasm Invasiveness; Neoplasm Metastasis; Prostatic Neoplasms
PubMed: 21474778
DOI: 10.1073/pnas.1102808108 -
Analytical Chemistry Aug 2018Protein catalyzed capture agents (PCCs) are synthetic antibody surrogates that can target a wide variety of biologically relevant proteins. As a step toward developing a...
Protein catalyzed capture agents (PCCs) are synthetic antibody surrogates that can target a wide variety of biologically relevant proteins. As a step toward developing a high-throughput PCC pipeline, we report on the preparation of a barcoded rapid assay platform for the analysis of hits from PCC library screens. The platform is constructed by first surface patterning a micrometer scale barcode composed of orthogonal ssDNA strands onto a glass slide. The slide is then partitioned into microwells, each of which contains multiple copies of the full barcode. Biotinylated candidate PCCs from a click screen are assembled onto the barcode stripes using a complementary ssDNA-encoded cysteine-modified streptavidin library. This platform was employed to evaluate candidate PCC ligands identified from an epitope targeted in situ click screen against the two conserved allosteric switch regions of the Kirsten rat sarcoma (KRas) protein. A single microchip was utilized for the simultaneous evaluation of 15 PCC candidate fractions under more than a dozen different assay conditions. The platform also permitted more than a 10-fold savings in time and a more than 100-fold reduction in biological and chemical reagents relative to traditional multiwell plate assays. The best ligand was shown to exhibit an in vitro inhibition constant (IC) of ∼24 μM.
Topics: Allosteric Regulation; Allosteric Site; Biotinylation; DNA, Single-Stranded; Drug Evaluation, Preclinical; Enzyme Inhibitors; Humans; Microarray Analysis; Proto-Oncogene Proteins p21(ras); Streptavidin
PubMed: 29979578
DOI: 10.1021/acs.analchem.8b00706 -
Nucleic Acids Research May 2000As a trial practical application, we have applied optimized microfabricated electrophoresis devices, combined with enzymatic mutation detection methods, to the...
As a trial practical application, we have applied optimized microfabricated electrophoresis devices, combined with enzymatic mutation detection methods, to the determination of single nucleotide polymorphism (SNP) sites in the p53 suppressor gene. Using clinical samples, we have achieved robust assays with quality factors as good as conventional electrophoresis in approximately 100 s. This is 10 and 50 times faster than capillary and slab gel electro-phoresis, respectively. The method was highly accurate with an average error of mutation site measurement of only +/-5 bp. No clean-up of the digestion mixtures was needed prior to injection. This greatly simplifies sample handling relative to capillary instruments, which is important for high-throughput screening applications. Following identification, absolute mutation determination of the screened samples was achieved in a second microdevice optimized for four-color DNA sequencing. Total run time was 25 min in this second device and sequencing data were in full agreement with ABI Prism 377 sequencing runs which required 3.5 h. The tandem application of microdevices for location then full characterization of SNPs appears to confirm many of the improvements claimed for future application of microdevices in practical scaled screening for mutational analysis.
Topics: DNA Mutational Analysis; DNA, Complementary; Electrophoresis, Polyacrylamide Gel; Endodeoxyribonucleases; Genes, p53; Humans; In Vitro Techniques; Miniaturization; Oligonucleotide Array Sequence Analysis; Polymorphism, Single Nucleotide; Reproducibility of Results; Sequence Analysis, DNA
PubMed: 10756210
DOI: 10.1093/nar/28.9.e43 -
Analytical Sciences : the International... Dec 2021We present a resin-packed microchannel that can reduce the radiation exposure risk and secondary radioactive wastes during uranium (U) separation by downscaling the...
We present a resin-packed microchannel that can reduce the radiation exposure risk and secondary radioactive wastes during uranium (U) separation by downscaling the separation using a microchip. Two types of microchips were designed to densely pack the microchannels with resins. The microchannels had almost the same cross-sectional area, but different outer circumferences. A satisfactory separation performance could be obtained by arranging more than ca. 10 resins along the depth and width of the microchannels. A resin-packed microchannel is an effective separation technique for determining the U concentration via inductively coupled plasma mass spectrometry owing to its ability to avoid the contamination of equipment by cesium, and to reduce the matrix effect. The size of the separation site was scaled down to <1/5000 compared to commonly used counterparts. The radiation exposure risk and secondary radioactive wastes can be reduced by 10- and 800-fold, respectively, using a resin-packed microchannel.
Topics: Anion Exchange Resins; Cesium; Seawater; Uranium
PubMed: 34305053
DOI: 10.2116/analsci.21P110 -
Sensors (Basel, Switzerland) Aug 2016The remarkable advantages micro-chip platforms offer over cumbersome, time-consuming equipment currently in use for bio-analysis are well documented. In this research, a...
The remarkable advantages micro-chip platforms offer over cumbersome, time-consuming equipment currently in use for bio-analysis are well documented. In this research, a micro-chip that includes a unique magnetic actuator (MA) for the manipulation of superparamagnetic beads (SPBs), and a magnetoresistive sensor for the detection of SPBs is presented. A design methodology, which takes into account the magnetic volume of SPBs, diffusion and heat transfer phenomena, is presented with the aid of numerical analysis to optimize the parameters of the MA. The MA was employed as a magnetic flux generator and experimental analysis with commercially available COMPEL™ and Dynabeads(®) demonstrated the ability of the MA to precisely transport a small number of SPBs over long distances and concentrate SPBs to a sensing site for detection. Moreover, the velocities of COMPEL™ and Dynabead(®) SPBs were correlated to their magnetic volumes and were in good agreement with numerical model predictions. We found that 2.8 μm Dynabeads(®) travel faster, and can be attracted to a magnetic source from a longer distance, than 6.2 μm COMPEL™ beads at magnetic flux magnitudes of less than 10 mT. The micro-chip system could easily be integrated with electronic circuitry and microfluidic functions, paving the way for an on-chip biomolecule quantification device.
Topics: Biosensing Techniques; Equipment Design; Lab-On-A-Chip Devices; Magnetics; Microspheres; Numerical Analysis, Computer-Assisted; Polystyrenes
PubMed: 27571084
DOI: 10.3390/s16091369 -
Nucleic Acids Research 2006Miniaturized integrated DNA analysis systems have largely been based on a multi-chamber design with microfluidic control to process the sample sequentially from one...
Miniaturized integrated DNA analysis systems have largely been based on a multi-chamber design with microfluidic control to process the sample sequentially from one module to another. This microchip design in connection with optics involved hinders the deployment of this technology for point-of-care applications. In this work, we demonstrate the implementation of sample preparation, DNA amplification, and electrochemical detection in a single silicon and glass-based microchamber and its application for the multiplexed detection of Escherichia coli and Bacillus subtilis cells. The microdevice has a thin-film heater and temperature sensor patterned on the silicon substrate. An array of indium tin oxide (ITO) electrodes was constructed within the microchamber as the transduction element. Oligonucleotide probes specific to the target amplicons are individually positioned at each ITO surface by electrochemical copolymerization of pyrrole and pyrrole-probe conjugate. These immobilized probes were stable to the thermal cycling process and were highly selective. The DNA-based identification of the two model pathogens involved a number of steps including a thermal lysis step, magnetic particle-based isolation of the target genomes, asymmetric PCR, and electrochemical sequence-specific detection using silver-enhanced gold nanoparticles. The microchamber platform described here offers a cost-effective and sample-to-answer technology for on-site monitoring of multiple pathogens.
Topics: Bacillus subtilis; Bacteria; DNA, Bacterial; Electrodes; Escherichia coli; Glass; Gold; Nanoparticles; Oligonucleotide Array Sequence Analysis; Oligonucleotide Probes; Point-of-Care Systems; Polymerase Chain Reaction; Silicon
PubMed: 17000638
DOI: 10.1093/nar/gkl702 -
Frontiers in Immunology 2015T cells are pivotal in the immune defense against cancers and infectious agents. To mount an effector response against cancer cells, T cells need to migrate to the...
T cells are pivotal in the immune defense against cancers and infectious agents. To mount an effector response against cancer cells, T cells need to migrate to the cancer-site, engage in contacts with cancer cells, and perform their effector functions. Adoptive T cell therapy is an effective strategy as treatment of complications such as relapse or opportunistic infections after hematopoietic stem cell transplantations. This requires a sufficient amount of cells that are able to expand and respond to tumor or viral antigens. The cytokines interleukin (IL)-2 and IL-7 drive T cell differentiation, proliferation, and survival and are commonly used to expand T cells ex vivo. Here, we have used microchip-based live-cell imaging to follow the migration of individual T cells, their interactions with allogeneic monocytes, cell division, and apoptosis for extended periods of time; something that cannot be achieved by commonly used methods. Our data indicate that cells grown in IL-7 + IL-2 had similar migration and contact dynamics as cells grown in IL-2 alone. However, the addition of IL-7 decreased cell death creating a more viable cell population, which should be beneficial when preparing cells for immunotherapy.
PubMed: 25972868
DOI: 10.3389/fimmu.2015.00196 -
Biomicrofluidics Mar 2015This paper describes a new and facile approach for the formation of pore-spanning bilayer lipid membranes (BLMs) within a poly(dimethylsiloxane) (PDMS) microfluidic...
This paper describes a new and facile approach for the formation of pore-spanning bilayer lipid membranes (BLMs) within a poly(dimethylsiloxane) (PDMS) microfluidic device. Commercially, readily available polycarbonate (PC) membranes are employed for the support of BLMs. PC sheets with 5 μm, 2 μm, and 0.4 μm pore diameters, respectively, are thermally bonded into a multilayer-stack, reducing the pore density of 0.4 μm-pore PC by a factor of 200. The BLMs on this support are considerably stable (a mean lifetime: 17 h). This multilayer-stack PC (MSPC) membrane is integrated into the PDMS chip by an epoxy bonding method developed to secure durable bonding under the use of organic solvents. The microchip has a special channel for guiding a micropipette in the proximity of the MSPC support. With this on-site injection technique, tens to hundreds of nanoliters of solutions can be directly dispensed to the support. Incorporating gramicidin ion channels into BLMs on the MSPC support has confirmed the formation of single BLMs, which is based on the observation from current signals of 20 pS conductance that is typical to single channel opening. Based on the bilayer capacitance (1.4 pF), about 15% of through pores across the MSPC membrane are estimated to be covered with BLMs.
PubMed: 26015832
DOI: 10.1063/1.4919066