Ivar Giaever

Applied BioPhysics, Inc · Owner

A personal account of the invention, growth and commercialization of Electric Cell-substrate Impedance Sensing (ECIS) by the inventors of the technology. From the first experiments at the General Electric Research and Development Center in the early 1980s to the outgrowth of applications and finally the incorporation of Applied BioPhysics, Inc., th...

In this study adherent animal cells were grown to confluence on circular gold-film electrodes of 250 μm diameter that had been deposited on the surface of a regular culture dish. The impedance of the cell-covered electrode was measured at designated frequencies to monitor the behavior of the cells with time. This approach is referred to as electric...

Some times unlikely events happens: How can a mechanical engineer from Norway end up with a Nobel Prize in Physics? I had the great fortune to receive the prize in Physics for using electron tunneling to measure the energy gap in superconductors. In this talk I will recollect some of the events that led to this discovery and hopefully be able to co...

Living mammalian cells can be perturbed by high pressure in a continuous and controlled manner. The effects can be easily measured and quantified using the well-established method of Electric Cell-substrate Impedance Sensing (ECIS). ECIS is an electrical biosensor that quantitatively monitors behaviours of living cells including spreading and adhes...

The main reason to start a business is probably to try to get rich, but our motivations were different (not that we mind making money). First it was and continues to be very difficult to get funding for interdisciplinary science in the USA. The regular granting agencies very often encourage research across the fields but fail to provide the money;...

Using an electrical measurement known as electric cell-substrate impedance sensing (ECIS), we have recorded the dynamics of viral infections in cell culture. With this technique, cells are cultured on small gold electrodes where the measured impedance mirrors changes in attachment and morphology of cultured cells. As the cells attach and spread on...

A new method combining optical and electrical impedance measurements is described that enables submicroscopic cell movements to be monitored. The cells are grown on small gold electrodes that are transparent to light. This modified electrical cell-substrate impedance sensor (ECIS) allows simultaneous microscopic recording of both growth and motilit...

Intercellular Ca2+ oscillations are a universal mode of signalling in both excitable and non-excitable cells. Here, we study the relationship between Ca2+ signalling and coherent changes in adhesion properties by measuring the transepithelial impedance across bradykinin-stimulated Madin-Darby canine kidney (MDCK) cell layers grown on a microelectro...

Confluent cell monolayers in tissue culture are fragile and can easily be mechanically disrupted, often leaving an area devoid of cells. This opening in the cell sheet is then repopulated, because the cells on the fringe of the damage, which are no longer contact-inhibited, move into the available space. This mechanical disruption is often done del...

The lumen of blood vessels is lined with a monolayer of endothelial cells (EC). In this work, electric cell-substrate impedance sensing (ECIS) was used to monitor the effect of fluid shear stress (FSS) on the morphology and function of cultured EC layers. Confluent layers of bovine aortic endothelial cells (BAEC) were grown on small gold electrodes...

In the present study we describe an electrical impedance analysis of BSC cells cultured on gold electrodes (250 im in diameter) that were modified with polypyrrole/heparin composites using electrochemical deposition. Atomic force microscope images show that the composite layer has a porous bulk structure and a very rough surface topology. An electr...

Here we present research detecting the invasive activities of metastatic cells in vitro using electric cell-substrate impedance sensing (ECIS). The assay is based on previous microscopic observations, where metastatic cells added over established endothelial cell layers were observed to attach to and invade the cell layer. Human umbilical vein endo...

Here we present research detecting the invasive activities of metastatic cells in vitro using electric cell-substrate impedance sensing (ECIS). The assay is based on previous microscopic observations, where metastatic cells added over established endothelial cell layers were observed to attach to and invade the cell layer. Human umbilical vein endo...

Here we describe various experiments that address the efficiency of loading extracellular probes into the cytoplasm of adherent mammalian cells (normal rat kidney, Madin-Darby canine kidney, and African green monkey) by means of in situ electroporation. Subsequent cell recovery from the electroporation pulse was monitored electrically in real time...

Electric cell-substrate impedance sensing (ECIS) has been developed as a non-invasive means to follow cell behavior in culture. In this method cells are cultured on small (250 micrometer diameter) gold film electrodes. The impedance of the electrode is measured by an AC current about 1 microampere. When challenged by biochemical or physical stimuli...

It has been said that Thomas Edison's greatest invention was that of the "Research Laboratory" as a social institution. My greatest discovery was when I learned at 29 years of age that it was possible to work in such an institution and get paid for doing research. I had become interested in physics, gotten a job at General Electric Research Laborat...

A flow chamber that allows the electric cell-substrate impedance sensing (ECIS) system to be used to evaluate the dynamic changes in endothelial electrical impedance during exposure to fluid flow was developed. The quantified changes in electrical resistance and capacitance were correlated to alterations in endothelial morphology and function known...

This talk addresses the electroporation efficiency when mammalian cells (NRK, MDCK, BSC) cultured on small gold-film electrodes, are electroporated. The elctroporation was done with sinusoidal AC voltage pulses at varying frequency, amplitude and duration. Permeability and reclosure of the plasma membrane was evaluated by the uptake into the cytopl...

It has long been realized that fibroblastic and epithelial cells establish recognizable patterns in tissue culture. This behavior implies that neighboring cells interact with one another to produce organized populations. Interaction between cells that are separated by many intervening cells is also possible and is demonstrated here using a special...

This article describes the optimization of an experimental technique referred to as electric cell-substrate impedance sensing (ECIS) to monitor attachment and spreading of mammalian cells quantitatively and in real time. The method is based on measuring changes in AC impedance of small gold-film electrodes deposited on a culture dish and used as gr...

To determine how histamine regulates endothelial barrier function through an integrative cytoskeletal network, we mathematically modeled the resistance across an endothelial cell-covered electrode as a function of cell-cell, cell-matrix, and transcellular resistances. Based on this approach, histamine initiated a rapid decrease in transendothelial...

When mammalian cells are seeded out in tissue culture they will drift down to the substratum, attach firmly and spread onto a pre-adsorbed protein layer and not directly to the naked substrate. Normally the protein layer will be a mixture of proteins obtained from serum added to the tissue culture medium, but it is also possible to pre-adsorb pure...

When mammalian cells are seeded out in tissue culture they will drift down to the surface and firmly attach. The cells attach to a preadsorbed protein layers and not to the naked surface. Normally the protein layer will be a mixture of protein obtained from serum added to the tissue culture medium, but it is also possible to preadsorb pure protein...

Great advances have been made in quantifying biochemical and physiological activities in cultured cells. It has, however, been difficult to quantify changes of cell morphology. A method has now been developed that can continuously and non-invasively track morphological changes of adherent cells and provide quantitative data from both sparse and con...

It has previously been demonstrated that oscillations occur in actively growing yeast cultures. These oscillations occur because yeast cells synchronize their glycolytic pathway following a saturation period. Periodic changes in the levels of intermediate metabolites in glycolysis as well as changes in pH ofthe media have been measured, that demons...

Transepithelial resistance (TER) measurement has often been used to study the paracellular transport properties of epithelia grown on permeable filters, especially the barrier function of tight junctions. However, the TER value includes another source, the resistance caused by cell-substrate contact, that may give rise to a high TER value if cell-s...

The manipulation force microscope is a novel atomic force microscope adapted to measuring the force necessary to displace micron-size samples adhering to surfaces. It has successfully characterized the adhesion of both proteins and living cells to substrates. This instrument enables measurement of samples not previously accessible to atomic force m...

The adhesion forces of cervical carcinoma cells in tissue culture were measured by using the manipulation force microscope, a novel atomic force microscope. The forces were studied as a function of time and temperature for cells cultured on hydrophilic and hydrophobic polystyrene substrates with preadsorbed proteins. The cells attached faster and s...

Orbital fibroblasts exhibit a phenotype distinct from that of other types of fibroblasts. Addition of prostaglandin E2 (PGE2) to culture medium elicits a dramatic change in orbital fibroblast morphology. That response is mediated through the generation of cAMP. Orbital fibroblasts can generate high levels of PGE2 through induction by proinflammator...

The force acting between a protein molecule and a nonbiological surface is of great importance in biotechnology. We adsorb protein-covered microspheres on glass and polystyrene substrates in Tris-buffered saline. Using a novel atomic force microscopy technique, we show that proteins adsorb better on hydrophobic than hydrophilic surfaces and that th...

An in vitro toxicity assay based on an emerging biosensor technology referred to as ECIS (electric cell-substrate impedance sensing) has been developed and tested. In ECIS, adherent cells are cultured, on small gold electrodes and electrical impedance is monitored with a computer-interfaced instrument. Subtle changes in cell morphology and behavior...

Fibroblasts and epithelial cells organize themselves in distinct patterns in tissue culture which indicates that neighboring cells communicate. A striking example of such communication is the oscillatory behavior of Madin-Darby canine kidney (MDCK) cells reported here. These oscillations were discovered using a biosensor referred to as ECIS (Electr...

Previously, we demonstrated the role of nitric oxide (NO) in transforming epithelial cells from a stationary to locomoting phenotype [E. Noiri, T. Peresleni, N. Srivastava, P. Weber, W.F. Bahou, N. Peunova, and M. S. Goligorsky. Am. J. Physiol. 270 (Cell Physiol. 39): C794-C802, 1996] and its permissive function in endothelin-1-stimulated endotheli...

Many different rhythms or oscillations exists in biological systems; the circadian rhythm is probably the best known, but other oscillations are also common. The slimemold Physarum is a classical example of ultradian oscillations occurring in a single multinucleated cell, and the period is a few minutes long. It is also reasonable well known that o...

Endothelin (ET) synthesis is enhanced at sites of ischemia or in injured vessels. The purpose of this study was to explore the possibility of autocrine stimulation of endothelial cell migration by members of the endothelin family. Experiments with microvascular endothelial cell transmigration in a Boyden chemotactic apparatus showed that endothelin...

We examined the contribution of actin-myosin contraction to the modulation of human umbilical vein endothelial cell focal adhesion caused by histamine and thrombin. Focal adhesion was measured as the electrical resistance across a cultured monolayer grown on a microelectrode. Actin-myosin contraction was measured as isometric tension of cultured mo...

We have developed a new analytical measurement for anchored cells in tissue culture that can track morphological changes of adherent cells. In this method, referred to as electric cell-substrate impedance sensing (ECIS), cells are cultured on small gold film electrodes and the impedance is probed with a 1 micro Amp current at 4 kHz. In the past, we...

Transepithelial impedance of Madin-Darby canine kidney cell layers is measured by a new instrumental method, referred to as electric cell-substrate impedance sensing. In this method, cells are cultured on small evaporated gold electrodes, and the impedance is measured in the frequency range 20-50,000 Hz by a small probing current. A model for imped...

Orbital fibroblasts from patients with Graves' ophthalmopathy, when treated with prostaglandin E2 (PGE2), become stellate and develop prominent cellular processes. In this paper, we describe results of studies designed to characterize the action of PGE2 on orbital fibroblast shape changes in vitro. Orbital and dermal fibroblasts were incubated with...

Although many biosensors are designed to be highly specific for a particular analyte, there is interest in designing sensors that can respond to a wide variety of chemical and physical stimuli in an integrated fashion. A famous case of this before the advent of biotechnology was the practice of bringing a canary into a coal mine. Because of the bir...

Orbital fibroblasts from patients with Graves’ ophthalmopathy, when treated with prostaglandin E, (PGE,), become stellate and develop prominent cellular processes. In this paper, we describe results of studies designed to characterize the action of PGE, on orbital fibroblast shape changes in vitro. Orbital and dermal fibroblasts were incubated with...

A novel electrical method for measuring the contraction rhythm occurring in Physarum polycephalum is described. The plasmodium is cultured in a dish containing two electrically separated chambers, and when bridging the two chambers the plasmodial veins of the Physarum complete an electrical circuit. The method is well suited for in situ measurement...

Summary The rhythmic contraction pattern in plasmodia ofPhysarum polycephalum was studied to determine whether characteristic changes occur during the synchronized nuclear division. An electrical method that measures the contraction rhythm in situ during several cell cycles was used. Biopsies of the plasmodia were taken at 17 min intervals for prec...

It is common in animal cell culture to utilize media that are buffered with bicarbonate ions and elevated CO2 concentrations in the incubator gas. Many modern incubators maintain these concentrations by monitoring the level of CO2 and injecting the gas when the value drops below a certain set point. This feedback system results in small oscillation...

Advantage was taken of DNA transfection techniques to investigate the effect of the pSV2-neo plasmid and its derivatives on recipient NIH 3T3 cell motion. Cell spreading and motion were followed by a newly developed electrical method to monitor cell morphology, referred to as electric cell-substrate impedance sensing. Using this method, we found th...

Describes a "whole cell" biosensor that is very useful in the field of animal cell tissue culture and in many ways is akin to the canary in a coal mine. The heart of this method, called ECIS for electric cell-substrate impedance sensing, is a small gold electrode immersed in tissue culture medium. When cells attach and spread on this electrode, the...

Fibroblasts derived from distinct anatomical regions appear to differ in regard to their behavior in culture. These differences may reflect functions of these cells in vivo that are tissue specific. Moreover, intrinsic differences in fibroblasts may underlie the site-specific connective tissue manifestations associated with systemic disease. We hav...

A newly developed method of detecting morphology and morphological changes of adherent mammalian cells cultured on thin film gold electrodes (electrical cell-substrate impedance sensing) has been used to measure the response of cells to external electrical stimuli. When sufficiently high ac voltage pulses were applied, the impedance of the cell lay...

An electrical biosensor is described that can continuously track morphological changes of adherent cells providing quantitative data from both sparse and confluent cultures. The method is capable of detecting vertical motion of cells of the order of 1 nm, much below the resolution of an optical microscope.

Summary form only given as follows: Great advances have been made in quantifying biochemical and physiological activities in cultured cells. It has, however been difficult to quantify changes of cell morphology. A method has now been developed that can continuously and non-invasively track morphological changes of adherent cells and provide data fr...

When an electrical potential of order one volt is induced across a cell membrane for a fraction of a second, temporary breakdown of ordinary membrane functions may occur. One result of such a breakdown is that molecules normally excluded by the membrane can now enter the cells. This phenomenon, generally referred to as electropermeabilization, is k...

In the plasmodial stage the slime mold Physarum polycephalum exhibits oscillatory phenomena. The ectoplasm contracts regularly, resulting in oscillatory (shuttle) streaming of the endoplasm. This periodic phenomenon represents a biological clock that is not well understood. We have studied quantitatively the stability of the rhythmic contractions i...

An electrical resistance method was developed to measure volume changes in substratum-attached monolayer cultures. Astrocytes in primary monolayer cultures prepared from neonatal rat cerebral cortex were placed in a confined channel containing a balanced salt solution, and the electrical resistance of the channel was measured using an applied alter...

Cell movements in confluent fibroblastic cell layers have been followed with a new instrumental method. Using the electric cell-substrate impedance sensor (ECIS), cell motions on the scale of nanometers are manifested as fluctuations in the impedance of small gold film electrodes that serve as cell substrata. These fluctuations have been recorded a...

We have developed an electrical method to study endothelial cell shape changes in real time in order to examine the mechanisms of alterations in the endothelial barrier function. Endothelial shape changes were quantified by using a monolayer of endothelial cells grown on a small (10(-3) cm2) evaporated gold electrode and measuring the changes in el...

A new electrical assay to measure the attachment and spreading of cells in tissue culture has been developed and substantiated by comparison with a more conventional assay. Small gold electrodes are vacuum deposited on the bottom of standard polystyrene culture dishes and coated with various proteins. As mammalian fibroblasts attach and spread on t...

Motility is a fundamental property of mammalian cells that normally is observed in tissue culture by time lapse microscopy where resolution is limited by the wavelength of light. This paper examines a powerful electrical technique by which cell motion is quantitatively measured at the nanometer level. In this method, the cells are cultured on small...

Cell spreading and cell locomotion arise from forces exerted by actin microfilaments upon the substratum. Using modified protein films at fluorocarbon oil--water interfaces as substrates, we have measured some minimal mechanical properties required of these films to support cell spreading forces in vitro. For murine 3T3-L1 fibroblasts, complete cel...

A novel type of whole-cell biosensor referred to as a cell-substrate impedance sensor (CIS) that electrically monitors the motion and morphology of anchored mammalian cells is discussed. In the sensor, cells are cultured on small gold electrodes and their attachment, spreading, and locomotions are detected as impedance changes. These measurements c...

A new method to monitor macrophage attachment on protein-coated surfaces and spreading in response to activating agents is described. Murine macrophages were cultured on small gold electrodes coated with protein, and attachment and spreading were detected as electrical impedance changes. The rate of attachment of cells to fibronectin-coated electro...

Hurst's rescaled range analysis has been applied to electrical signals that reflect the motion of fibroblastic human cells. According to the analysis the cells have a fractional Brownian motion with a Hurst exponent of about H=0.65. The normal WI-38 cell line has a Hurst exponent that is slightly smaller than the transformed WI-38 VA13 cell line.

A simple technique is described that is suitable for rapid screening of hybridoma microculture fluids for monoclonal antibody producing hybrids. The procedure measures increase in light scatter due to the antigen-antibody reaction on a surface of indium metal coated upon glass and does not require use of a labeled second antibody. Techniques minimi...

Everything we are is determined by the genetic information we carry. Forays into conquering disease and birth defects depend on understanding genes first. Through the millenia man has known that certain characteristics can be inherited. However, just what and where these genes were remained a mystery until the experiments of Avery, MacLeod, and McC...

Dispersed fluorocarbon oil droplets were used as the antigen-carrier particle in an immunological agglutination assay. The extent of agglutination was found to vary with the percentage of antigen to that of inert protein used in the solution to coat and stabilize the oil droplets. Maximum sensitivity was observed when 6.25% of the protein adsorbed...

Mammalian cells can be cultured and therefore studied in vitro. Normally, the cells' morphology and other static properties are observed with the aid of a light microscope. A method is described here that allows observation of the dynamical aspects of cultured cells. Mammalian fibroblasts are cultured in polystyrene dishes that contain evaporated g...

Mammalian fibroblasts have been cultured on evaporated gold electrodes subjected to an alternating electric field at 4000 Hz. The system consists of a large (approximately equal to 2 cm2) and a small (approximately equal to 3 X 10(-4) cm2) electrode bathed in tissue culture medium. The applied electric field produces a voltage drop at the boundary...

This solid-phase immunoassay for detection of rheumatoid factor involves a specially prepared indium surface that renders protein layers visible and therefore does not require tagged protein. Using appropriate discriminatory values, we found agreement with results of a standard latex-fixation test in controls and patients with rheumatoid arthritis....

Anchorage-dependent fibroblasts can be cultured by using as a substrate the protein layer that spontaneously forms at the liquid-liquid interface between fluorocarbon fluids and tissue culture medium. For this novel substrate to be effective in supporting confluent cell layers, the protein monolayer must support the stresses exerted by spreading fi...

Anchorage-dependent cell growth is demonstrated on microcarriers of fluorocarbon fluid formed by emulsification and stabilized with polylysine.

We have cultured the murine cell lines 3T3-L1 and SV-T2 using as a substrate the layer of denatured protein that forms at the phase boundary between culture medium and fluorocarbon fluids. The growth patterns observed on these interfaces differ from those seen on conventional solid substrates. Depending on the cell strain and the composition of the...

A simple aqueous deposition procedure is described which can produce partial monolayer coverages of ferritin on field-emitter tips. By employing the same deposition procedure with other biomolecules, it may now be possible to prepare field-emitter tips with known molecular coverages — a prerequisite for any attempt to image biomolecules with field-...

We examined the suitability of a rapid immunological technique to determine the amount of aspartate aminotransferase (EC 2.6.1.1) isoenzymes in human serum or tissue extracts. Purified isoenzymes were absorbed as a monolayer to the surface of an indium metal film on glass. The enzyme retains immunological reactivity, allowing the specific binding o...

A device is described which is used to measure the concentration of antigen in solution. A wedge-shaped gap containing the antigen to be detected is bounded on one side by a reservoir containing specific antiserum and on the opposite side by a detection slide which has been previously coated with a layer of adsorbed antigen. As the antibodies diffu...

The adsorption of ferritin molecules onto both Lexan polycarbonate and carbon surfaces has been studied with the help of an electron microscope. The amount of ferritin adsorbed is a sensitive function of both the pH and the salinity of the ferritin solution. Complete coverage of a surface could be obtained only under very special conditions. Comple...

Protein deposition on field-emitter tips has been examined using Transmission Electron Microscopy to view the protein coated tip profile. A single layer of adsorbed protein is barely if at all detectable, but double and triple layers produced by the immunologic reaction can be directly observed. As a result, the thickness and morphology of antigen-...

A novel method of radioimmune assay is described. It departs from conventional assays in that the radioiodine is reacted with protein following, rather than before, the antigen-antibody reaction. Using a BSA/anti-BSA system, the assay can detect antigen concentrations down to approximately 100 ng/ml.

When most mammalian cells are propagated in tissue culture, they are attached to a solid surface. This surface is commonly covered with a layer of serum protein. In this study we looked at the effect of the protein layer on the interaction between the cells and the surface by precoating the surface with various pure protein molecules. We found no d...

A new general immunology test is described which is capable of detecting protein in the 1 microgram/ml range. First an antigen is adsorbed in a small area of a glass surface. The glass is exposed to an antibody solution and the antibody will attach specifically to the antigen. Next the slide is washed, dried and sprayed lightly with plastic particl...

An attempt is made to detect carcinoembryonic antigen (CEA) at clinically interesting concentrations by using a simple immunologic surface test. Antibodies to CEA are detected in a direct test at concentrations below 1 ng/ml. The sensitivity of this assay is mainly limited by diffusion to the reacting surface. CEA is detected in an inhibition test...

Part of the immunology system in vertebrates deals with the highly specific reaction between a foreign protein (antigen) and the proteins manufactured as a response (antibodies). The immunology reaction will take place in vitro, and it is used in the medical field to identify specific proteins. This lecture will discuss the antibody-antigen reactio...

When the hepatitis B antigen is first adsorbed on a specially prepared metal surface, the subsequent reaction with its specific antibody produces complexes that are visible to the unaided eye. Application of this principle has led to the development of tests for both antigen and antibody that compare favorably with the sensitivity obtained by radio...

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