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At least 1,171 records · Page 65Linked to original sources
PubMed2001

Analysis of single-cell differences by use of an on-chip microculture system and optical trapping.

A method is described for continuous observation of isolated single cells that enables genetically identical cells to be compared; it uses an on-chip microculture system and optical tweezers. Photolithography is used to construct microchambers with 5-microm-high walls made of thick photoresist (SU-8) on the surface of a glass slide. These microchambers are connected by a channel through which cells are transported, by means of optical tweezers, from a cultivation microchamber to an analysis microchamber, or from the analysis microchamber to a waste microchamber. The microchambers are covered with a semi-permeable membrane to separate them from nutrient medium circulating through a "cover chamber" above. Differential analysis of isolated direct descendants of single cells showed that this system could be used to compare genetically identical cells under contamination-free conditions. It should thus help in the clarification of heterogeneous phenomena, for example unequal cell division and cell differentiation.

Bacteria↗
PubMed2001

Optical DNA-sensor chip for real-time detection of hybridization events.

An optical sensor system based on evanescent field excitation of fluorophore-labeled DNA-targets specifically binding to immobilized DNA probes has been developed, thus enabling for real-time analysis of hybridization events. Oligonucleotide probes are directly immobilized on the surface of the disposable sensor chip via biotin/neutravidin linkage and hybridize to complementary Cy5-labeled target DNA in the sample; this is recorded as an increase in the fluorescence signal. Under optimized conditions the hybridization rate was constant and directly proportional to the target concentration. When an 18mer oligonucleotide was used as a probe a linear calibration curve was obtained for a 56mer single-stranded DNA target derived from the neomycin phosphotransferase gene, a selection marker in a variety of genetically modified plants, with an estimated lower limit of detection of 0.21 nmol L(-1). No cross-hybridization to a 51mer actin DNA target was observed and even a single-nucleotide mismatch led to a negligible signal. A shutter in the readout device enabled separate detection of targets hybridizing to probes immobilized at the inlet and outlet sides, respectively, of the flow channel. This opens a route toward a real-time DNA array format with analysis times as short as 1-2 min. As a realistic sample a Cy5-labeled 56 bp PCR product was measured after separation of the double-stranded DNA by simple heat denaturation with a detection limit clearly lower than that of traditional gel electrophoresis.

Actins↗
PubMed2005

Low-volume amplification on chemically structured chips using the PowerPlex16 DNA amplification kit.

In forensic DNA analysis, improvement of DNA typing technologies has always been an issue. It has been shown that DNA amplification in low volumes is a suitable way to enhance the sensitivity and efficiency of amplification. In this study, DNA amplification was performed on a flat, chemically structured glass slide in 1-microl reaction volumes from cell line DNA contents between 1,000 and 4 pg. On-chip DNA amplification reproducibly yielded full allelic profiles from as little as 32 pg of template DNA. Applicability on the simultaneous amplification of 15 short tandem repeats and of a segment of the Amelogenin gene, which are routinely used in forensic DNA analysis, is shown. The results are compared to conventional in-tube amplification carried out in 25-microl reaction volumes.

Case-Control Studies↗
PubMed2001

Fly-like visuomotor responses of a robot using aVLSI motion-sensitive chips.

We explore the use of continuous-time analog very-large-scale-integrated (aVLSI) neuromorphic visual preprocessors together with a robotic platform in generating bio-inspired behaviors. Both the aVLSI motion sensors and the robot behaviors described in this work are inspired by the motion computation in the fly visual system and two different fly behaviors. In most robotic systems, the visual information comes from serially scanned imagers. This restricts the form of computation of the visual image and slows down the input rate to the controller system of the robot, hence increasing the reaction time of the robot. These aVLSI neuromorphic sensors reduce the computational load and power consumption of the robot, thus making it possible to explore continuous-time visuomotor control systems that react in real-time to the environment. The motion sensor provides two outputs: one for the preferred direction and the other for the null direction. These motion outputs are created from the aggregation of six elementary motion detectors that implement a variant of Reichardt's correlation algorithm. The four analog continuous-time outputs from the motion chips go to the control system on the robot which generates a mixture of two behaviors--course stabilization and fixation--from the outputs of these sensors. Since there are only four outputs, the amount of information transmitted to the controller is reduced (as compared to using a CCD sensor), and the reaction time of the robot is greatly decreased. In this work, the robot samples the motion sensors every 3.3 ms during the behavioral experiments.

Animals↗
PubMed2005

Dynamic analysis of metabolic effects of chloroacetaldehyde and cytochalasin B on tumor cells using bioelectronic sensor chips.

PURPOSE: To study the interplay of drugs and energy metabolism of tumor cells, metabolic changes induced by chloroacetaldehyde and cytochalasin B were analyzed in colon carcinoma cells LS174T. METHODS: O(2)-consumption and extracellular acidification were recorded using a bioelectronic sensor-chip system, which monitors these parameters in a culture continuously for at least 24 h. In parallel cultures cell number, cellular ATP-content, mitochondrial transmembrane potential, and the content of reactive oxygen species (ROS) were determined. RESULTS: When cell death was induced by chloroacetaldehyde (50 muM), the rate of acidification declined gradually for the next 15 h, while O(2)-consumption decreased rapidly within 30 min. This correlated with a loss in mitochondrial potential. However, cellular ATP-level showed a transient increase at 2 h; also ROS levels increased up to 6 h. In cells treated with cytochalasin B (2 muM), which inhibits glucose uptake, the rate of O(2)-consumption increased and the acidification activity dropped, even upon glutamine depletion. Mitochondrial membrane potential transiently increased after 1 h, while ATP-content decreased; there was no change in the level of ROS. CONCLUSION: The pattern of changes in basic energy metabolism differs with the type of cell death and growth inhibition involved in the cytotoxic action of two different drugs.

Acetaldehyde↗
PubMed2006

A hybrid poly(dimethylsiloxane) microsystem for on-chip whole blood filtration optimized for steroid screening.

Miniaturized biochemical devices in glass, silicon and polymer materials are starting to find their way from the academic laboratories to real-life applications. However, most attention has been given to miniaturize the downstream functions of various microfluidic systems, leaving the sample introduction and preparation steps to more conventional, bulkier solutions. For point-of-care diagnostics in particular, it becomes crucial to be able to handle complex human samples in a miniaturized format.In this work, we report on a microsystem for on-chip sample preparation that is able to remove blood cells from whole blood. The hybrid system consists of a commercially available membrane filter incorporated into a poly(dimethylsiloxane) (PDMS) casted device. Membrane materials were evaluated on the bases of low nonspecific adsorption of free and protein-bound testosterone as analyte substance. The hybrid system including a hydrophilic polypropylene filter successfully removed blood cells from diluted human whole blood. Surface oxidation was sufficient to make the plasma filtrate flow through the membrane filter and the channel system by capillary force alone and thus no external pumping source was needed.

Biocompatible Materials↗
PubMed2006

Sensitivity enhancement for colorimetric glucose assays on whole blood by on-chip beam-guidance.

In this paper, we present a novel concept for optical beam-guidance to significantly enhance the sensitivity of colorimetric assays by extending the optical path length through the detection cell which linearly impacts the resulting attenuation of a probe beam according to the law of Beer-Lambert. In our setup, the incident probe beam is deflected by 90( composite function) into the chip plane at monolithically integrated V-grooves to pass a flat detection cell at its full width (i.e., with a path length of 10 mm) instead of its usually much smaller height. Afterwards, the attenuated beam is redirected by another V-groove towards an external detector. The general beam-guidance concept is demonstrated by a glucose assay on human whole blood on a centrifugal microfluidic "lab-on-a-disk" platform made of COC. We achieve an excellent linearity with a correlation coefficient (R (2)) of 0.997 paired with a lower limit of detection (200 microM) and a good reproducibility with a coefficient of variation (CV) of 4.0% over nearly three orders of magnitude. With an accelerated sedimentation of cellular constituents by centrifugal forces, the sample of whole blood can be analyzed in a fully integrated fashion within 210 s. This time-to-result can even be improved by the numerical extrapolation of the saturation value. Additionally, the direct assay on whole blood also shows a negligible correlation with the hematocrit of the blood sample.

Blood Chemical Analysis↗
PubMed1983

The effect of the filling material in the tissue reactions following apical plugging of the root canal with dentin chips. A histologic study in monkeys' teeth.

The object of the present work was to observe whether the root canal filling material has some influence in the periapical healing process after apical plugging with dentin chips. Monkeys' teeth were filled with nine different filling materials in use today. Ninety days after the treatment, the histologic data showed no differences in the results with the nine materials studied. This fact suggests that the filling material does not influence the healing process when the plug is present.

Animals↗
PubMed1996

DNA chips: analysing sequence by hybridization to oligonucleotides on a large scale.

Complete sequences of genomes and comprehensive sets of cDNA sequences open the way to a hugh range of biological problems. There is a need for analytical methods that can deal with the large number of sequences in the data banks and, ideally, we would like to analyse all sequences together. Gel-based sequencing is a serial process that analyses one sequence at a time. Oligonucleotide arrays, or 'DNA chips', are miniature, parallel analytical devices, which could bring to sequence analysis and molecular genetics many of the advantages that semiconductor devices brought to computing.

Base Sequence↗
PubMed1994

Chip fractures of the triquetrum. Mechanism, classification and results.

Of 231 patients with fractures of the triquetrum, 65 were followed up after a mean period of 46.8 months. The fractures were classified and radiological and clinical follow-up was carried out. Pathomechanical aspects of the origin of the fracture were also considered. It appears that the chisel action of the dorso-proximal edge of the hamate striking against the fully extended and ulnar-deviated wrist is the major cause of the dorsal chip fracture of the triquetrum. Conservative therapy involving immobilization of the injured wrist for 3 weeks proved to be successful. If the bone fragment healed, it took 6 to 8 weeks. There was no indication for surgical intervention. We found no signs of post-traumatic instability of the joint. There was good vascularization of the bone, which excluded the possibility of avascular necrosis. All fractures involving the main body of the triquetrum united.

Carpal Bones↗
PubMed1996

Burn injuries caused by chip-pan fires: the Edinburgh experience.

Thirty-two patients were admitted to the South-East Scotland Burn Unit over a 4-year period (1990-94), with burns sustained as a consequence of chip-pan fires. This represented 7 per cent of all admissions to the unit. Fourteen patients came from the Edinburgh city area, and 18 from surrounding countries. There were 14 males and 18 females, and an average age of 51.6 years. The average burn surface area was 4.7 per cent. Eighteen patients had burns to their hands, this being the most common area involved. Fourteen patients were managed conservatively, and 18 needed operative intervention. The patients spent an average of 19.4 days in hospital, and they required 46.3 days for complete healing to take place. Prevention by increased community awareness and widespread education is necessary to minimize the incidence, morbidity and cost of this relatively common and preventable type of burn.

Adolescent↗
PubMed2004

Chip-based gel electrophoresis method for the quantification of half-antibody species in IgG4 and their by- and degradation products.

The inter-heavy-chain disulfide bonds of the IgG4 subclass can be described as being at equilibrium with the intra-chain disulfide bonds. This means that a fraction of IgG4 has noncovalently linked heavy chains (half-antibody). The percentage of half-antibodies produced depends upon the expression system used. Nondenaturing assays fail to separate the half-antibodies from the native form because two half-molecules are held together by noncovalent forces. The pharmaceutical industry needs a reliable denaturing assay for checking batch-to-batch consistency. Until now sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) has been the standard method used to detect the presence of half-antibodies. However, this technique is laborious and cannot be automated. Furthermore, cumbersome densitometric measurements are necessary for quantification. To overcome these disadvantages a chip-based gel electrophoresis method was investigated. In the nonreduced format the separation profile is compared with that from SDS-PAGE. The limit of quantification as a percentage of the amount applied, repeatability, reproducibility, and linearity are compared with those of SDS-PAGE. The amounts of half-antibody and of by- and degradation products were determined for several batches by using area percentage and by external calibration with IgG4 as a reference standard. Both methods allow avoidance of error introduction for the quantification as is the case by application of myosin as reference concentration. Both sets of results are compared with each other and with the results from SDS-PAGE. In the reduced format it is noted that the reduction of the inter-heavy-chain disulfide bridges proceeds faster than the reduction of the heavy-light-chain bonds. Therefore optimized conditions are necessary to obtain a complete reduction.

Antibodies↗
PubMed2004

On-chip micro-flow polystyrene bead-based immunoassay for quantitative detection of tacrolimus (FK506).

Tacrolimus (FK506) is a widely used immunosuppressant for preventing allograft rejection and the treatment of atopic dermatitis. FK506 necessitates therapeutic drug monitoring because of inter- and intrapatient variability and the lack of correlation between the administered dose and the blood concentration. Previous immunoassay-based methods required a relatively long assay time and troublesome liquid-handling procedures. In the present study, we aimed to establish a rapid monitoring method for FK506 determination by using a poly(dimethylsiloxane) (PDMS)-based microfluidic device. Polystyrene beads were coated with mouse anti-FK506 antibody and placed in the flow channel. As a competitive assay, sample solution was allowed to react in the flow channel. After the addition of the fluorogenic substrate, the fluorescent signal was observed under a microscope. As a result, the developed assay allowed a short detection time of approximately 15 min per each sample and a high sensitivity even by using only a single bead. The feasibility of performing a competitive assay using a PDMS-based antibody chip gives promising results over the existing immunoassay-based methods.

Animals↗
PubMed2005

Using surface plasmon resonance to directly identify molecules in a tripeptide library that bind tightly to a vancomycin chip.

This paper describes a procedure, based on direct binding, for identifying tight-binding ligands for a receptor immobilized on a sensor chip from an array of equimolar tripeptides using surface plasmon resonance. Vancomycin and a library of 96 tripeptides, with molecular weight ranging from 316 to 560 Da, were used as a model system to illustrate the procedure. A consensus structure of the strongest interacting peptides consisted of D-Ala at the C terminus and aromatic amino acid in the penultimate position. Ligands having this structure bound more tightly to vancomycin than the known D-Ala-D-Ala peptide. The throughput of our continuous assay is 96 compounds in 3.3 h, and the sample consumption is less than 2 microg per peptide and 1 ng for vancomycin. This procedure should be applicable to peptide libraries of greater complexity than that used here and to mixtures of small organic compounds.

Ligands↗
PubMed2006

Quantitative evaluation of cell-to-cell communication effects in cell group class using on-chip individual-cell-based cultivation system.

Cell-to-cell communication is considered to underlie the coordinated behavior and the multicellularity of cell group class, which cannot be explained only by the knowledge of lower class of life system from molecule to individual cell, because they are determined by at least two different ways: diffusible chemical signals and their direct physical contacts. We show in this paper a new method of individual-cell-based cell observation that can estimate the role of cell-to-cell communication, diffusible chemical signals, and physical contacts as separated properties, by applying an on-chip individual-cell-based cultivation system. The exchange of stationary phase medium on isolated individual Escherichia coli from exponential phase medium and the control of physical contacts indicated that the cell-to-cell direct contact did not affect the growth rate; only the communication through diffusible signals affects the growth rates as Hill's equation manner.

Cell Communication↗
PubMed2006

Quantitative measurement of damage caused by 1064-nm wavelength optical trapping of Escherichia coli cells using on-chip single cell cultivation system.

We quantitatively examined the possible damage to the growth and cell division ability of Escherichia coli caused by 1064-nm optical trapping. Using the synchronous behavior of two sister E. coli cells, the growth and interdivision times between those two cells, one of which was trapped by optical tweezers, the other was not irradiated, were compared using an on-chip single cell cultivation system. Cell growth stopped during the optical trapping period, even with the smallest irradiated power on the trapped cells. Moreover, the damage to the cell's growth and interdivision period was proportional to the total irradiated energy (work) on the cell, i.e., irradiation time multiplied by irradiation power. The division ability was more easily affected by a smaller energy, 0.36 J, which was 30% smaller than the energy that adversely affected growth, 0.54 J. The results indicate that the damage caused by optical trapping can be estimated from the total energy applied to cells, and furthermore, that the use of optical trapping for manipulating cells might cause damage to cell division and growth mechanisms, even at wavelengths under 1064 nm, if the total irradiation energy is excessive.

Cell Culture Techniques↗
PubMed2005

An inverted microcontact printing method on topographically structured polystyrene chips for arrayed micro-3-D culturing of single cells.

With the goal to investigate the relation of shape and function of single cells or clusters of cells in a 3-dimensional (3-D) microenvironment, we present a novel platform technology to create arrays of microwells on polystyrene (PS) chips for hosting cells in a local microenvironment characterized by controlled shape and surface chemistry. The micro-3-D cell culturing combines 2-dimensional chemical patterning with topographical microstructuring presenting to the cells a local 3-D host structure. Microwells of controlled dimensions were produced by a two-step replication process, based on standard microfabrication of Si, replica molding into poly(dimethylsiloxane), and hot embossing of PS. This allowed the production of large numbers of microstructured surfaces with high reproducibility and fidelity of replication. Using inverted micro contact printing, the plateau surface between the microwells was successfully passivated to block adsorption of proteins and prevent cell attachment by transfer of a graft-copolymer, poly(l-lysine)-g-poly(ethylene glycol). The surface inside the microwells was subsequently modified by spontaneous adsorption of proteins or functionalized PLL-g-PEG/PEG-X (X=biotin or specific, cell-interactive peptide) to elicit specific responses inside the wells. Preliminary cell experiments demonstrated the functionality of such a device to host single epithelial cells (MDCK II) inside the functionalized microwells and thus to control their 3-D shape. This novel platform is useful for fundamental cell-biological studies and applications in the area of cell-based sensing.

Adsorption↗
PubMed2006

Assessment of GaN chips for culturing cerebellar granule neurons.

In this work, the behaviors of cerebellar granule neurons prepared from 7-day-old Wistar rats on gallium nitride (GaN) were investigated. We believe that this is the first time that the GaN has been used as a substrate for neuron cultures to examine its effect on cell response in vitro. The GaN surface structure and its relationship with cells were examined by atomic force microscopy (AFM), metallography microscopy, scanning electron microscopy (SEM), lactate dehydrogenase (LDH) release and Western blot analysis. GaN is a so-called III-V compound semiconductor material with a wide bandgap and a relatively high bandgap voltage. Compared with silicon used for most neural chips, neurons seeded on GaN were able to form an extensive neuritic network and expressed very high levels of GAP-43 coincident with the neurite outgrowth. Therefore, the GaN structure may spatially mediate cellular response that can promote neuronal cell attachment, differentiation and neuritic growth. The favorable biocompatibility characteristics of GaN can be used to measure electric signals from networks of neuronal cells in culture to make it a possible candidate for use in a microelectrode array.

Animals↗