Abstract: The present invention relates to the use of porous silicon in the delivery of substances into cells. The porous silicon can be formed into micropiercers, microneedles and biolistic bullets for peenetration of the cell. The control of the pore size and porosity of the porous silicon allows tuning of the bioactivity of the porous silicon. The porous silicon is also resorbable and is therefore resorbed from the cells without leaving any particles or being seen as a foreign body. The present invention also relates to the methods of manufacturing the porous silicon micropiercers, microneedles, microelectrodes, biolistic bullets, and precipitation of calcium phosphate on a bioactive substrate, and their advantages over known methods of delivering materials into cells.

Abstract: Systems, methods and apparatus for stabilizing resistance measurements in electroporation systems so as to achieve high-accuracy measurements, and for controlling the pulse duration, or time constant, of electroporation pulses. Circuitry is provided to accurately determine the resistance of a sample in the electroporation system, for example a sample provided in an electroporation cuvette. Additionally, a resistance control system is provided to automatically measure the resistance of a sample, determine a capacitance and determine a parallel add-on resistance that substantially provides a desired pulse duration (time constant) for an electroporation pulse.

Abstract: Method and apparatus for accelerating micro particles for use in delivering DNA or a solid drug in which a shockwave is generated by applying a short pulse energy to a surface of a metal foil to be absorbed and cause vaporization and plasmatization of the metal foil. A jet is generated by a sudden expansion of metal gas and thereby the shockwave is generated on a surface of an opposite side of the metal foil on which the micro particles are arranged.

Abstract: The invention relates to a novel circuit arrangement for electrotransfection or electrofusion, which enables the transportation of DNA and/or other biologically active molecules to the nucleus of higher eukaryotic cells or the fusion of cells, independent of cell division and with reduced cell mortality.

Abstract: Transmembrane potential measurement methods using cationic dyes, and anionic dyes are provided. Compositions of the cationic and anionic dyes and microfluidic systems which include the dyes and membranes are provided in conjunction with processing elements for transmembrane potential measurements.

Abstract: Techniques for streaming electroporation. A representative but non-limiting method includes: generating a spatially inhomogeneous electric field with a pair of electrodes and displacing the pair of electrodes and a sample relative to one other while the electric field is substantially constant in terms of magnitude so that the sample is displaced across electric field lines for a time sufficient to effect electroporation.

Abstract: A detection chip capable of detecting large amounts of genes with high sensitivity is provided. The gene detection chip comprises a plurality of pins 12 as measurement electrodes, a common electrode 22 as a counter electrode therefor, and a tabular member 14 with a plurality of pin holes 15 for accommodating the pins. An arrangement may be adopted in which the diameters of the pin holes 15 taper off in the direction in which the pins are inserted, and the pins are held in place in the narrowest sections of the pin holes 15.

Abstract: The invention relates to an apparatus for and method of introducing a substance into an object, particularly into a cell or cellular material. In a preferred arrangement apparatus receives the cell and preferably locates it between first and second electrodes and applies a voltage pulse to cause a disruption in the cell wall. This causes the cell to become permeable. The substance may then be introduced, for example under a fluid pressure. Cells may then be inspected and sorted into transfected and non-transfected types. This may be achieved automatically, for example by using electrophoresis.

Abstract: Cuvette inserts adapted and configured to fit within an electroporation cuvette. The inserts each include a support structure that holds a porous membrane. When positioned within the cuvette, the membrane is positioned proximal the cuvette electrodes to facilitate membrane-based fusion of cells. In certain aspects, a tube extends through the support structure to allow for application of negative pressure in a convenient location away from electrode contacts and other components of the cuvette or cuvette holder.

Abstract: The invention provides apparatus and methods for subsecond lysis of selected cells in a cell chamber using a voltage pulse of 10 ms to 10 &mgr;s in duration followed by nearly simultaneous collection of the lysed cellular contents into a capillary electrophoresis tube or other suitable micro-collection device. Cell chambers and capillary electrophoresis tubes configured with electrodes for performing the electrical lysis are described. The influence of variables that govern the rate of cell lysis, such as inter-electrode distance, pulse duration, and pulse strength are also described. The methods are illustrated using fluorophores that are loaded into a cell then collected following electrical lysis, separated by electrophoresis, and then detected by laser-induced fluorescence detection in a capillary electrophoresis system.

Abstract: The present invention relates to a method and an apparatus for separating substances from or in biological materials.

Abstract: A shocking chamber for performing electroporation is constructed in the form of a base and a hinged lid which when lowered forms an enclosure with the base that fully encloses a cuvette and impresses a high voltage across the cuvette, the voltage connection becoming disengaged upon the simple raising of the lid. The base contains two pairs of electrical leads, one pair engaging the cuvette with spring-loaded contacts that provide electrical connections to the cuvette while helping to secure the cuvette in place, and the other pair joined to high-voltage terminals. A shunt built into the lid bridges the two pairs of leads in the base when the lid is closed and pivots out of the way to clear all leads when the lid is opened. These and other features of the construction provide the user with a safe and secure means of forming the high-voltage electrical connections used in electroporation in an apparatus that can be operated with one hand.

Abstract: A housing has a chamber containing a low speed, e.g., 6 rpm, rotating shaft of highly polished stainless steel to avoid bubbles and arcing. A fluid containing cells to be electroporated and molecules associated with the electroporation are introduced into the chamber formed by the gap between the shaft and housing in either batch or continuous flow modes. A capacitor network applies alternating positive and negative pulses of high voltage across the electrodes formed by the housing and shaft to electroporate the cells. After a few pulses, a resistive load is placed across the network to reduce the value of the pulses applied to the electrodes for a relatively longer period than the high voltage pulses to a relatively low harmless level.

Abstract: Drug candidate screening methods are applied to discover compounds with activity against ion channel targets. The method may include modulating the transmembrane potential of host cells in a plurality of sample wells with a repetitive application of electric fields so as to set the transmembrane potential to a level corresponding to a pre-selected voltage dependent state of a target ion channel.

Abstract: A carrier for gene detection as a means for prediction before treatment whether interferon therapy is valid or not for a patient, a method for detection of interferon therapy for an individual, an apparatus for gene detection, and a kit for detection of validity of interferon therapy.

Abstract: A method and apparatus for analyzing nucleic acids includes immobilizing nucleic probes at specific sites within a microchannel structure and moving target nucleic acids into proximity to the probes in order to allow hybridization and fluorescence detection of specific target sequences.

Abstract: The invention relates to an apparatus for and method of introducing a substance into an object, particularly into a cell or cellular material. In a preferred arrangement apparatus receives the cell and preferably locates it between first and second electrodes and applies a voltage pulse to cause a disruption in the cell wall. This causes the cell to become permeable. The substance may then be introduced, for example under a fluid pressure. Cells may then be inspected and sorted into transfected and non-transfected types. This may be achieved automatically, for example by using electrophoresis.

Abstract: For sample picking on a cryosubstrate, on which multiple cryopreserved samples are each positioned at preselected sample positions, individual samples are selectively separated mechanically or thermally from the cryosubstrate and transferred to a target substrate.

Abstract: Delivery of metal particles to living tissue, then applying external energy that interacts with the metal particles, is found to selectively increase the energy deposition and interaction surrounding the metal particles. The method is useful to improve treatment of various conditions, since targeted cells may be selectively altered or killed. Metal particles are also loaded into cells or membrane vesicles by placing metal seed particles into the cells or vesicles, then chemically depositing additional metal on the metal seed particles. The metal particles are useful to improve imaging and therapies by their interaction with externally applied energy.

Abstract: An apparatus and method are disclosed for in vitro transformation of living cells. The apparatus, which is formed as a microelectromechanical device by surface micromachining, can be used to temporarily disrupt the cell walls or membrane of host cells one at a time so that a particular substance (e.g. a molecular tag, nucleic acid, bacteria, virus etc.) can be introduced into the cell. Disruption of the integrity of the host cells (i.e. poration) can be performed mechanically or electrically, or by both while the host cells are contained within a flow channel. Mechanical poration is possible using a moveable member which has a pointed or serrated edge and which is driven by an electrostatic actuator to abrade, impact or penetrate the host cell. Electroporation is produced by generating a relatively high electric field across the host cell when the host cell is located in the flow channel between a pair of electrodes having a voltage applied therebetween.

Abstract: Electroporation is performed in a controlled manner in either individual or multiple biological cells or biological tissue by monitoring the electrical impedance, defined herein as the ratio of current to voltage in the electroporation cell. The impedance detects the onset of electroporation in the biological cell(s), and this information is used to control the intensity and duration of the voltage to assure that electroporation has occurred without destroying the cell(s). This is applicable to electroporation in general. In addition, a particular method and apparatus are disclosed in which electroporation and/or mass transfer across a cell membrane are accomplished by securing a cell across an opening in a barrier between two chambers such that the cell closes the opening. The barrier is either electrically insulating, impermeable to the solute, or both, depending on whether pore formation, diffusive transport of the solute across the membrane, or both are sought.

Abstract: A micromachined cell lysis device with electrodes that are spaced by less than 10 &mgr;m from one another. The cells are attracted to the space between the electrodes and then lysed.

Abstract: A shocking chamber for performing electroporation is constructed in the form of a base and a hinged lid which when lowered forms an enclosure with the base that fully encloses a cuvette and impresses a high voltage across the cuvette, the voltage connection becoming disengaged upon the simple raising of the lid. The base contains two pairs of electrical leads, one pair engaging the cuvette with spring-loaded contacts that provide electrical connections to the cuvette while helping to secure the cuvette in place, and the other pair joined to high-voltage terminals. A shunt built into the lid bridges the two pairs of leads in the base when the lid is closed and pivots out of the way to clear all leads when the lid is opened. These and other features of the construction provide the user with a safe and secure means of forming the high-voltage electrical connections used in electroporation in an apparatus that can be operated with one hand.

Abstract: The invention concerns a method for treating an aqueous flow colonized by cells with a pulsed electric field applied to a flow, characterized in that the applied field is substantially parallel to the direction of flow and to its application to the transfer of nucleic acids (RNA, DNA, oligonucleotides) into cells, to the transfer of proteins to cells, to the extraction of cytoplasmic macromolecules and molecules contained in the cells, to cell fusion and the production of hybrids and/or to insertion of membrane proteins. It also concerns an electropulsing chamber, a method for destroying cells and a membrane permeabilization method.

Abstract: The invention provides methods and compositions for combinatorially decoding arrays.

Abstract: An improved electrode for use in generating an electrical field in a saline solution is provided. In particular, a continuous crystalline metal nitride coated electrode is provided for use in a variety of saline solution applications, such as in an electrophoresis device for separating proteins or nucleic acids or an electroporation apparatus for the encapsulation of biologically-active substances in various cell populations. A method and apparatus are provided for the encapsulation of biologically-active substances in red blood cells, characterized by an optionally automated, continuous-flow, self-contained electroporation system which allows withdrawal of blood from a patient, separation of red blood cells, encapsulation of a biologically-active substances in the cells, and optional recombination of blood plasma and the modified red blood cells thereby producing blood with modified biological characteristics.

Abstract: Methods for determining the presence of double stranded nucleic acids in a sample are provided. In the subject methods, nucleic acids present in a fluid sample are translocated through a nanopore, e.g. by application of an electric field to the fluid sample. The current amplitude through the nanopore is monitored during the translocation process and changes in the amplitude are related to the passage of single- or double-stranded molecules through the nanopore. The subject methods find use in a variety of applications in which the detection of the presence of double-stranded nucleic acids in a sample is desired, e.g. in hybridization assays, such as Northern blot assays, Southern blot assays, array based hybridization assays, etc.

Abstract: A solid state device is formed through thin film deposition techniques which results in a self-supporting thin film layer that can have a precisely defined channel bored therethrough. The device is useful in the chacterization of polymer molecules by measuring changes in various electrical characteristics as molecules pass through the channel. To form the device, a thin film layer having various patterns of electrically conductive leads are formed on a silicon substrate. Using standard lithography techniques, a relatively large or micro-scale aperture is bored through the silicon substrate which in turn exposes a portion of the thin film layer. This process does not affect the thin film. Subsequently, a high precision material removal process is used (such as a focused ion beam) to bore a precise nano-scale aperture through the thin film layer that coincides with the removed section of the silicon substrate.

Abstract: Integrated microfluidic devices comprising at least an enrichment channel (10) and a main electrophoretic flowpath (12) are provided. In the subject integrated devices, the enrichment channel and the main electrophoretic flowpath are positioned so that waste fluid flows away from said main electrophoretic flowpath through a discharge outlet (6). The subject devices find use in a variety of electrophoretic applications, including clinical assays, high throughput screening for genomics and pharmaceutical applications, point-or-care in vitro diagnostics, molecular genetic analysis and nucleic acid diagnostics, cell separations, and bioresearch generally.

Abstract: An apparatus for electrically contacting biological cells suspended in a liquid has a substrate having at least one opening and an electrode for electrically contacting a cell immobilized above the opening. A contact unit with a contact tip is arranged below said opening. A top end of the contact tip projects into the opening in such a way that it comes to bear against a cell membrane of the immobilized cell. The contact tip has a contact channel which ends at its top end. A hydrodynamic low pressure can be exerted upon the cell membrane via the contact channel, and the electrode is electrically connected to the contact channel. In a method for electrically contacting biological cells suspended in a liquid, a cell is immobilized above an opening provided in a substrate, and the immobilized cell is contacted via at least one electrode. For contacting the immobilized cell, a hydrodynamic low pressure is generated acting on a cell membrane through a contact tip projecting into the opening.

Abstract: A device for determining the characteristics of a biomass, includes: a probe (S,S′) including electrodes, two or four in number, for injecting a current into a medium containing biological cells and for reading the voltage applied to the medium, and a resistance for measuring the injected current; a conditioning unit (10) including an alternating voltage generator (112) and a circuit (200) processing current and voltage measuring signals, so as to deliver signals (C,G) respectively of measurement of the capacity and conductance of the medium. The conditioning unit (10) includes a zero method measuring bridge (150) automatically controlled to deliver the signals (C,G) of measurement of the medium capacity and conductance. The invention is useful for characterizing media containing biological cells, in particular for controlling biological fermentation process.

Abstract: An array is placed in contact with or close proximity with an organ or tissue surface to cause electroporation for gene, protein, drug delivery in both ex vivo and in vivo applications. A low DC voltage with a short pulse duration, and long burst pulse is applied to the array. A long rest period is provided between pulse bursts to allow for cell recovery. To enable the application of a low voltage shock in a large organ with same transfection efficiency, four gene, protein and drug delivery systems are illustrated.

Abstract: High frequency interfacing to biochemical membranes, such as supported bilayers and cell membranes, is carried out by supporting a biochemical membrane on a support surface while allowing access to the surface of the membrane through an opening in the support. A sharp tipped probe is positioned adjacent to the exposed surface of the membrane. The probe may have an inner core tip and a coaxial shield around the core tip that is electrically insulated therefrom. Radio frequency power is supplied to the probe to apply a localized radio frequency field to the membrane adjacent to the probe. Transport and binding events at the membrane are detected by changes in the field transmitted through the membrane and received by a receiving probe, or reflected from the membrane and received by the transmitting probe, and coupled therefrom to a detector for detection.

Abstract: Cuvette inserts adapted and configured to fit within an electroporation cuvette. The inserts each include a support structure that holds a porous membrane. When positioned within the cuvette, the membrane is positioned proximal the cuvette electrodes to facilitate membrane-based fusion of cells. In certain aspects, a tube extends through the support structure to allow for application of negative pressure in a convenient location away from electrode contacts and other components of the cuvette or cuvette holder.

Abstract: A mass biosensor uses an intermediate avidin layer to facilitate binding of a biotinylated antibody to a measurement surface of the biosensor. The avidin layer can be added by the manufacturer of the biosensor, while the biotinylated layer can be added by the user. This two-phase method of chemically modifying the measurement surface significantly reduces the user time required to customize the measurement surface to render it capable of binding selected compounds. An organosilane coupling agent attached to the surface provides sites to which avidin is bound. Avidin acts as a universal receptor of biotinylated compounds with specific binding affinities. Biotinylated antibodies or other biotinylated compounds are added and bind to the immobilized avidin. Surface adsorption is reduced by washing the modified surface with biotin to block potential sites of weak bond formation, electrostatic and hydrophobic interactions.

Abstract: A molecule introducing apparatus is provided with a power supply circuit 20 for supplying power, a step-up transformer 30 which is supplied with power from the power supply circuit 20 and which outputs a high voltage, a switching transistor 22 that blocks off and allows the supply of power from the power supply circuit 20 to the step-up transformer 30 and which generates an instantaneous high voltage, and a pair of electrode probes 50 for applying the instantaneous high voltage generated at the step-up transformer 30 to a predetermined region in a body. The molecule introducing apparatus oscillates DNA arranged outside a cell by the instantaneous high voltage so as to introduce the DNA into the cell.

Abstract: Microchip devices are provided which include a substrate having a plurality of reservoirs containing a secondary device, a reacting component, or a combination thereof. At least one barrier layer covers each reservoir to isolate the reservoir contents from one or more environmental components outside the reservoirs. The barrier layer can be selectively disintegrated or permeabilized to expose the isolated contents to the one or more environmental components. The secondary device preferably includes a sensor or sensing component, for example, a biosensor, or a light detection or imaging device, such as an optical fiber. Preferred reacting components include catalysts and reagent, which may be immobilized in the reservoirs.

Abstract: The present invention relates to methods and apparatus for the encapsulation of biologically-active substances in various cell populations. More particularly, the present invention relates to a method and apparatus for the encapsulation of biologically-active substances in various cell populations in blood by electroporation to achieve therapeutically desirable changes in the physical characteristics of the various cell populations in blood.

Abstract: This system provides methods and apparatus for performing microanalytic and microsynthetic analyses and procedures. Specifically, the system provides a microsystem platform for use with a micromanipulation device to manipulate the platform by rotation, thereby utilizing the centripetal force resulting from rotation of the platform to motivate fluid movement through microchannels embedded in the microplatform. The microsystem platforms of the system are also provided having microfluidics components, resistive heating elements, temperature sensing elements, mixing structures, capillary and sacrificial valves, and methods for using these microsystems platforms for performing biological, enzymatic, immunological and chemical assays. An electronic spindle designed rotor capable of transferring electrical signals to and from the microsystem platforms of the system is also provided.

Abstract: The invention relates to a method for detecting analytes and to a device for carrying out the method, for use for analysis or diagnosis in the fields of chemistry, biochemistry, molecular genetics, food chemistry, biotechnology, the environment and medicine. Marker particles (5) with different electrical properties or a different relative permeability to those of the measuring solution (3) surrounding them are used to detect the analytes (8). The marker particles (5) either bond specifically to the analytes (8) or to a base (2) in competition with the analyte. The analytes (8) are detected by the changes in an electrical field or an electrical current generated by electrodes (2) or in an electrical voltage applied to an electrode or in a magnetic field, said changes being caused by marker particles which have bonded with the analytes or by marker particles which have instead bonded to the base in an electrical field.

Abstract: The present invention relates to methods and apparatus for the encapsulation of biologically-active substances in various cell populations. More particularly, the present invention relates to a method and apparatus for the encapsulation of biologically-active substances in various cell populations in blood by electroporation to achieve therapeutically desirable changes in the physical characteristics of the various cell populations in blood.

Abstract: A novel method and device for transporting and/or monitoring a fluid in a multi-port device 400, 800, 1000 used in a microfluidic system is provided. The multi-port device includes a substrate having a novel channel configuration. A first channel region 413 having a first port and a second port for transporting fluid therebetween is defined in the substrate. A second channel region 421 having a first port and a second port for applying electric current for heating fluid or for monitoring a fluid parameter therebetween is also defined in the substrate. In some embodiments, the first channel intersects 407 with the second channel. The heating or monitoring aspect of the invention can be used with a variety of biological reactions such as PCR, LCR, and others.

Abstract: An apparatus for automatically measuring minute membrane potential, based on a technique developed for controlling a membrane denaturation reaction without using a physical shearing force, for example, a method of causing the destruction of membrane at a limited portion of a living membrane by making a stimulus, such as light and a compound activated by the stimulus react with each other in a membrane, such as a living membrane, this method being applied to a minute electrode to facilitate the insertion thereof into a cell, which has been difficult in the use of a minute metal electrode, and enable membrane potential in a cell to be measured easily, the minute metal electrode enabling the integration thereof and the development of a neural interface in the barrier-free technology.

Abstract: A method for facilitating a delivery of a molecule into an interior space of a cell includes the steps of introducing a molecule into a target tissue comprising a cell and applying a substantially continuous low-level electric field to the target tissue. The field is applied for a duration sufficient to effect a change in porosity the cell of the target tissue sufficient to facilitate an entry of a desired molecule into an interior of the cell.

Abstract: A gas permeable cover for a cell culture or medium storing container which comprises a solid support sized to fit the opening of the container to be covered and has at least one hole which is covered with a permeable section. The permeable section is sufficiently permeable to oxygen and carbon dioxide that live cells can be sustained, and is substantially impermeable to water and water vapor such that the covered containers can be stored in non-humidified chambers without substantial evaporation of water. The cover further has a seal portion which forms a water-and gas-tight seal between the cover and the container when the cover is placed upon the container as configured for use.

Abstract: This relates to methods for the detection of psychoactive compounds in an in vitro neuronal tissue sample by detecting oscillations of extracellular voltage desirably before and after the introduction of a candidate sample onto an in vitro neuronal tissue sample and for devices useful in practicing the methods. Analysis of the extracellular voltage parameters leads to indication of the presence of psychoactive material in the candidate sample and information as to its pharmacological activity and/or composition. Further, it relates to a process of initiating and maintaining the presence of repetitive neuronal activity within the in vitro sample. Additionally, this includes a method for the stimulation of or initiation of repetitive neuronal activity, e.g.

Abstract: The invention comprises an apparatus and a method of effecting localized electroporation in a relatively small target area and for introducing foreign matter into cellular material in which the target area is located. The cellular material may be in vitro, in ovo or in vivo.

Abstract: Apparatus and methods are provided for interacting light with particles, including but not limited to biological matter such as cells, in unique and highly useful ways. Optophoresis consists of subjecting particles to various optical forces, especially optical gradient forces, and more particularly moving optical gradient forces, so as to obtain useful results. In biology, this technology represents a practical approach to probing the inner workings of a living cell, preferably without any dyes, labels or other markers.

Abstract: The invention concerns a device and a method for ultrasensitive detection comprising an electrically inert porous sheet sandwiched between two positively charged fine electrodes. The invention uses the capacity of the electrodes, resulting from their powerful electrostatic properties, for sensing marked antibody complexes related to antigens detected in the sample, and to leave marked antibodies non-complexed with said antigens free. The method consists in depositing a sample droplet on the porous material sheet upstream of the electrodes and in detecting downstream of the electrodes, in a signal-zone, the possible presence of an antigen-antibody complex which has diffused in the porous material. The invention is useful for detecting some millipicogrammes of antigen per milliliter of sample.

Abstract: A method for identifying a microorganism is described that includes abstracting gas or vapor associated with the microorganism from a detection region and flowing the same over an array of sensors of which an electrical property varies according to exposure to gases or vapors and observing the response of the sensors. An apparatus for detecting a microorganism is also disclosed having a detector means for detecting a gas or vapor associated with the microorganism which includes an array of sensors of which an electrical property varies according to exposure to the gases or vapors.