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At least 1,639 records · Page 91Linked to original sources
PubMed2003

Anisotropic stretch-induced hypertrophy in neonatal ventricular myocytes micropatterned on deformable elastomers.

Because cell shape and alignment, cell-matrix adhesion, and cell-cell contact can all affect growth, and because mechanical strains in vivo are multiaxial and anisotropic, we developed an in vitro system for engineering aligned, rod-shaped, neonatal cardiac myocyte cultures. Photolithographic and microfluidic techniques were used to micropattern extracellular matrices in parallel lines on deformable silicone elastomers. Confluent, elongated, aligned myocytes were produced by varying the micropattern line width and collagen density. An elliptical cell stretcher applied 2:1 anisotropic strain statically to the elastic substrate, with the axis of greatest stretch (10%) either parallel or transverse to the myofibrils. After 24 h, the principal strain parallel to myocytes did not significantly alter myofibril accumulation or expression of atrial natriuretic factor (ANF), connexin-43 (Cx-43), or N-cadherin (by indirect immunofluorescent antibody labeling and immunoblotting) compared with unstretched controls. In contrast, 10% transverse principal strain resulted in continuous staining of actin filaments (rhodamine phalloidin); increased immunofluorescent labeling of ANF, Cx-43, and N-cadherin; and upregulation of protein signal intensity by western blotting. By using microfabrication and microfluidics to control cell shape and alignment on an elastic substrate, we found greater effects for transverse than for longitudinal stretch in regulating sarcomere organization, hypertrophy, and cell-to-cell junctions.

Actins↗
PubMed2003

Compression of biocompatible liquid-filled HSA-alginate capsules: determination of the membrane mechanical properties.

Compression experiments between two parallel plates are performed on a series of biocompatible HSA-alginate capsules with two different membrane thicknesses. The capsule geometry and size as well as the average membrane thickness are first measured. The compression set-up is fitted with a sensitive force transducer that allows measurement of the compression force as a function of plate separation. The response of the capsule is analyzed by assuming different constitutive models for the membrane, where the shear and surface dilatation effects are accounted. An apparent area dilatation modulus is then computed for different values of the plate separation and required to remain constant as the capsule deformation increases. This allows identification of plausible constitutive laws for the membrane material.

Alginates↗
PubMed2003

Levitation and movement of human tumor cells using a printed circuit board device based on software-controlled dielectrophoresis.

In this study we describe an original, efficient, and innovative printed circuit board (PCB) device able to generate dielectrophoresis-based, software-controlled cages that can be moved to any place inside a microchamber. Depending on their dielectrophoretic properties, eukaryotic cells can be "entrapped" in cages and moved under software control. The main conclusion gathered from the experimental data reported is that the PCB device based on dielectrophoresis permits levitation and movement of different tumor cells at different dielectrophoresis conditions. The results presented herein are therefore the basis for experiments aimed at forced interactions or separation of eukaryotic cells using "lab-on-a-chip." In fact, because many cages can be controlled at the same time, and two or more cages can be forced to share the same or a different location, it is possible, in principle, either to bring in contact cells of a differing histotype or to separate them.

Animals↗
PubMed2005

Positioning, displacement, and localization of cells using ultrasonic forces.

This paper presents a method and a device to position and displace cells. The cells are suspended in a fluid layer trapped between the device and an arbitrary surface such as an object slide or a wafer. The device vibrates at ultrasonic frequencies causing a pressure field in the fluid layer. This pressure field results in a force-field capable of positioning cells. Depending on the way in which the device is excited a 2-D or 3-D force-field can be generated, positioning the cells in lines or points respectively. Furthermore, it is possible to subsequently displace the cells with micrometer accuracy. This has been demonstrated using HL60 and MCF10A cells, and can be achieved without causing damage to the cells.

Biophysical Phenomena↗
PubMed2007

Degradation of supercoiled plasmid DNA within a capillary device.

Supercoiled plasmid DNA is susceptible to fluid stress in large-scale manufacturing processes. A capillary device was used to generate controlled shear conditions and the effects of different stresses on plasmid DNA structure were investigated. Computational fluid dynamics (CFD) analysis was employed to characterize the flow environment in the capillary device and different analytical techniques were used to quantify the DNA breakage. It was found that the degradation of plasmid DNA occurred at the entrance of the capillary and that the shear stress within the capillary did not affect the DNA structure. The degradation rate of plasmids was well correlated with the average elongational strain rate or the pressure drop at the entrance region. The conclusion may also be drawn that laminar shear stress does not play a significant role in plasmid DNA degradation.

Computer Simulation↗
PubMed2007

Cell patterning using magnetite nanoparticles and magnetic force.

Technologies for fabricating functional tissue architectures by patterning cells precisely are highly desirable for tissue engineering. Although several cell patterning methods such as microcontact printing and lithography have been developed, these methods require specialized surfaces to be used as substrates, the fabrication of which is time consuming. In the present study, we demonstrated a simple and rapid cell patterning technique, using magnetite nanoparticles and magnetic force, which enables us to allocate cells on arbitrary surfaces. Magnetite cationic liposomes (MCLs) developed in our previous study were used to magnetically label the target cells. When steel plates placed on a magnet were positioned under a cell culture surface, the magnetically labeled cells lined on the surface where the steel plate was positioned. Patterned lines of single cells were achieved by adjusting the number of cells seeded, and complex cell patterns (curved, parallel, or crossing patterns) were successfully fabricated. Since cell patterning using magnetic force may not limit the property of culture surfaces, human umbilical vein endothelial cells (HUVECs) were patterned on Matrigel, thereby forming patterned capillaries. These results suggest that the novel cell patterning methodology, which uses MCLs, is a promising approach for tissue engineering and studying cell-cell interactions in vitro.

Cell Culture Techniques↗
PubMed2004

Ciliary reorientation is evoked by a rise in calcium level over the entire cilium.

Internal Ca2+ levels control the pattern of ciliary and flagellar beating in eukaryotes. In ciliates, ciliary reversal is induced by a rise in intra-ciliary Ca2+, but the mechanism by which Ca2+ induces reversal is not known. We injected the fluorescent Ca2+ indicator Calcium Green into a ciliate Didinium nasutum and observed the intra-ciliary Ca2+ level during the initial reversed stroke preceding spontaneous cyclic reversed beating. In D. nasutum, Ca2+ rose throughout the length of the cilia undergoing initial reversed stroke. Electron microscopy with a combined oxalate-pyroantimonate method showed Ca2+ deposits distributed throughout the reversed cilia. We injected caged Ca2+ into D. nasutum and irradiated the base or mid region of the cilia with UV to locally increase Ca2+ concentration. Uncaging Ca2+ in the middle of the cilia produced reversal distally, but not proximally to the site of Ca2+ release. These results strongly suggest that not only Ca2+ influx sites, but also Ca2+ binding sites and vectoral bending machineries for ciliary reversal, are distributed throughout the cilium.

Animals↗
PubMed1992

Mechanical perturbation of webbed edges in 3T3 cells.

We have previously described actin edge-bundles (AEBs) as cables of microfilaments lining the webbed edges of 3T3 cells (Zand and Albrecht-Buehler: Cell Motil. Cytoskeleton 13:195-211, 1989). We have suggested that AEBs, along with their cell-substratum adhesions, resist cortical tension and prevent the collapse of cytoplasm towards the nucleus. In this paper, we report several stages of AEB disassembly and re-formation induced by the following micro-manipulations: (1) Scoring of the webbed edge of a 3T3 cells with a micro-needle. As a result the sides of the score retracted and the severed AEB appeared to disassemble down to its terminal adhesion points. The retraction stopped after 20-40 seconds and the cells formed a webbed edge with large curvature. Over a period of 20-80 minutes, the new web decreased in length and depth, until it regained its approximate original shape. (2) Bending of cell processes at acute angles. As a result the processes moved until they projected at right angles to the side of the cell and formed new webs gradually expanded their area. In both cases, the nascent webs were lined by actin edge-bundles.

3T3 Cells↗
PubMed1995

Ni2+ inhibition induces asymmetry in axonemal functioning and bend initiation of bull sperm.

Bull sperm extracted with 0.1% Triton X-100 can be reactivated to full motility with 0.33 mM Mg-ATP (sperm models). When motile sperm models are treated with 0.66 mM NiSO4, spontaneous motility is lost. During the transition to motility arrest, the beat becomes progressively more asymmetric, finally arresting at one extreme of the beat cycle. After spontaneous motility has been lost, the flagellum retains the ability to respond to mechanical stimulation. If a microprobe is used to bend the flagellum in the direction opposite to its own prevailing curvature and released, the recoil is rapid and overshoots the equilibrium position. When the same flagellum is manipulated in the opposite direction (into a tighter bend of the existing curve), the recoil is slower and does not exceed the initial bend. If a microprobe is used to carefully bend the whole flagellum into a curve, the flagellum will resume continuous beating, but only if the imposed bend is in the direction opposite the natural curvature. The reinstated beating activity (mechanical reactivation) is sustained as long as the flagellum is held by the microprobe. The rate of change of the shear angle in these mechanically reactivated, Ni(2+)-inhibited sperm suggests an impaired rate of sliding on one side of the axoneme compared to similarly restrained control sperm. It appears that Ni2+ has a selective inhibitory effect on the dynein arms that bend the flagellum in one direction. Furthermore, the remaining functional arms activate only when the flagellum is bent in the direction opposing their own action.

Animals↗
PubMed1995

Experimental manipulation of gamma-tubulin distribution in Arabidopsis using anti-microtubule drugs.

gamma-Tubulin-specific antibodies stain the microtubule (Mt) arrays of Arabidopsis suspension cells in a punctate or patchy manner. During division, staining of kinetochore fibers and the phragmoplast is extensive, except in the vicinity of the plus ends at the metaphase plate and cell plate. gamma-Tubulin localization responds to low levels of colchicine, with staining receding farther toward the minus (pole) ends of kinetochore fibers. At higher drug concentrations, gamma-tubulin also associates with abnormal Mt foci as well as with the surface of the daughter nuclei facing the phragmoplast. During UV-induced recovery from colchicine, gamma-tubulin increases along the presumptive minus ends of mitotic Mts as well as the phragmoplast near the daughter nuclei. With CIPC, immunostaining is concentrated around the centers of focal Mt arrays in multipolar spindles. In the presence of taxol, Mts are more prominent but the mitotic apparatus and phragmoplast are abnormal. As with CIPC, gamma-tubulin is concentrated at focal arrays. Increased punctate staining is also present in interphase arrays, with fluorescent dots often located at the ends of Mts. These results support a preferential association between gamma-tubulin and Mt minus ends, but are also consistent with more general binding along the walls of Mts. Thus, minus ends (and Mt nucleation sites) may be present throughout plant Mt arrays, but gamma-tubulin may also serve another function, such as in structural stabilization.

Amino Acid Sequence↗
PubMed1990

Personal computer interface and software for the ZONAX microscope attachment controller.

We describe the hardware and software of a general-purpose interface that permits a personal computer (PC) running MS-DOS to control cytometric devices, e.g., a scanning stage, shutters, and focus motor attached to a microscope. This note explains how to use the interface to convert the ZONAX microscope attachment controller from 8080-based hardware control to PC-based control.

Computer Peripherals↗
PubMed1991

Experimental and theoretical investigations on the validity of the geometrical optics model for calculating the stability of optical traps.

We have developed a computer program based on the geometrical optics approach proposed by Roosen to calculate the forces on dielectric spheres in focused laser beams. We have explicitly taken into account the polarization of the laser light and thd divergence of the laser beam. The model can be used to evaluate the stability of optical traps in a variety of different optical configurations. Our calculations explain the experimental observation by Ashkin that a stable single-beam optical trap, without the help of the gravitation force, can be obtained with a strongly divergent laser beam. Our calculations also predict a different trap stability in the directions orthogonal and parallel to the polarization direction of the incident light. Different experimental methods were used to test the predictions of the model for the gravity trap. A new method for measuring the radiation force along the beam axis in both the stable and instable regions is presented. Measurements of the radiation force on polystyrene spheres with diameters of 7.5 and 32 microns in a TEM00-mode laser beam showed a good qualitative correlation with the predictions and a slight quantitative difference. The validity of the geometrical approximations involved in the model will be discussed for spheres of different sizes and refractive indices.

Lasers↗
PubMed1991

Single beam optical trapping integrated in a confocal microscope for biological applications.

Confocal microscopy is very useful in biology because of its three dimensional imaging capacities and has proven to be an excellent tool to study the 3D organization of, for instance, cell structures. This property of confocal microscopy makes it also very suitable for observation during guidance of the three dimensional manipulation of single cells or cell elements. Therefore we decided to integrate a confocal microscope and a single beam optical manipulator into a single instrument. The advantage of optical manipulation over mechanical techniques is that it is non-invasive and therefore may be applied on living (micro-) organisms and cells. The creation of an effective single beam optical trap requires the use of a high numerical aperture (N.A.) objective to focus the laser beam. In this paper we briefly discuss the vertical or axial force exerted on a sphere in a single beam trap. The axial force on a sphere placed on the optical axis, caused by reflection and refraction, is calculated applying a electromagnetic vector diffraction theory to determine the field distribution in the focal region. One of the results is that the particle also experiences a vertical trapping force towards the focusing lens when it is in the strongly convergent part of the field in addition to the known negative signed trapping force in the divergent part of the field. Further we describe an instrumental approach to realize optical trapping in which the optical trap position is controlled by moving the focusing objective only.(ABSTRACT TRUNCATED AT 250 WORDS)

Algorithms↗
PubMed1991

Compliance of bacterial polyhooks measured with optical tweezers.

In earlier work, a single-beam gradient force optical trap ("optical tweezers") was used to measure the torsional compliance of flagella in wild-type cells of Escherichia coli that had been tethered to glass by a single flagellum. This compliance was nonlinear, exhibiting a torsionally soft phase up to 180 degrees, followed by a torsionally rigid phase for larger angles. Values for the torsional spring constant in the soft phase were substantially less than estimates based on the rigidity determined for isolated flagellar filaments. It was suggested that the soft phase might correspond to wind-up of the flagellar hook, and the rigid phase to wind-up of the stiffer filament. Here, we have measured the torsional compliance of flagella on cells of an E. coli strain that produces abnormally long hooks but no filaments. The small-angle compliance of these cells, as determined from the elastic rebound of the cell body after wind-up and release, was found to be the same as for wild-type cells. This confirms that the small-angle compliance of wild-type cells is dominated by the response of the hook. Hook flexibility is likely to play a useful role in stabilizing the flagellar bundle.

Bacterial Proteins↗
PubMed1991

Application of laser optical tweezers in immunology and molecular genetics.

Optical tweezers, based on a compact diode pumped Nd:YAG laser providing 350 mW at 1,064 nm coupled into a Zeiss IM 35 microscope, were used to sort CD4+ T cells into a capillary for further mechanical handling and to establish contact between single human natural killer (NK) cells and human erythroleukemia cells (K562) as targets. After contact and a lag phase of a few tens of seconds, the target cell starts to change its morphology and membrane blebbing occurs. The kinetics of the attack of the NK cell on K562 cells is not straightforward but governed by temporal oscillations in the shape of the target cell (zeosis). In a second application, the optical tweezers are combined with a UV laser microbeam based on a pulsed UV laser and with flow cytometry and sorting. With the pulsed laser, segments of sorted chromosome 1 of the chinese hamster karyotype (CHV 79) can be easily micro-dissected and subsequently collected using the optical tweezers. This allows preparation of a few hundred chromosome segments per day without mechanical contact and in an absolutely sterile way and thus may provide an interesting basic technique in any type of genome sequencing project.

Animals↗
PubMed1991

Laser induced cell fusion in combination with optical tweezers: the laser cell fusion trap.

A single-beam gradient force optical trap was combined with a pulsed UV laser microbeam in order to perform laser induced cell fusion. This combination offers the possibility to selectively fuse two single cells without critical chemical or electrical treatment. The optical trap was created by directing a Nd:YAG laser, at a wavelength of 1.06 microns, into a microscope and focusing the laser beam with a high numerical aperture objective. The UV laser microbeam, produced by a nitrogen-pumped dye laser (366 nm), was collinear with the trapping beam. Once inside the trap, two cells could be fused with several pulses of the UV laser microbeam, attenuated to an energy of approximately 1 microJ/pulse in the object plane. This method of laser induced cell fusion should provide increased selectivity and efficiency in generating viable hybrid cells.

Animals↗
PubMed1993

Optimizing a method of protein extraction for two-dimensional electrophoretic separation of proteins from planarians (Platyhelminthes, Turbellaria).

Different procedures for microscale extraction of proteins from small amounts of tissue of planarians (Platyhelminthes, Turbellaria) to be analyzed by two-dimensional gel electrophoresis (2-D PAGE) are compared. Three extraction methods were assessed: (i) extraction of soluble proteins with nondenaturing Tris buffers, (ii) extraction with Tris buffer containing the anionic detergent sodium dodecyl sulfate (SDS), and (iii) denaturing extraction under reducing conditions in the presence of urea and Nonidet P-40 (NP-40) with or without SDS. Buffers combining minute concentrations of SDS (0.01%), denaturing concentrations of urea (8M) and alkaline pH solubilized the greatest number of proteins without detectable proteolysis. Neither the presence of protease inhibitors nor higher concentrations of SDS improved protein extraction. We have applied this method to planarians to detect proteins specific to the pharynx. The resulting two-dimensional pattern shows a larger number of specific spots than in previous extraction methods.

Animals↗