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 features a composition for the delivery of bioactive agents into cells that includes a delivery matrix, an anionic or zwitterionic compound, and a bioactive agent, e.g. a peptide, protein, or nucleic acid. The compositions of the invention can be used to deliver bioactive compounds, such as nucleic acids encoding immunostimulatory peptides and/or therapeutic proteins.

Abstract: A method of highly efficiently transferring various selected molecules into various cells and a method of fusing cells. Cells and/or selected molecules such as polynucleotide are treated with cold gas plasma to thereby transfer the selected molecules located around cells into the cells, or cells are fused by treating the cells with cold gas plasma. Moreover, an apparatus for transferring selected molecules or fusing cells having a cold gas plasma generation unit for transferring selected molecules into cells is provided.

Abstract: An electrospraying apparatus and/or method is used to coat particles. For example, a flow including at least one liquid suspension may be provided through at least one opening at a spray dispenser end. The flow includes at least particles and a coating material. A spray of microdroplets suspending at least the particles is established forward of the spray dispenser end by creating a nonuniform electrical field between the spray dispenser end and an electrode electrically isolated therefrom. The particles are coated with at least a portion of the coating material as the microdroplet evaporates. For example, the suspension may include biological material particles.

Abstract: We describe a process for generating multilayer particles comprising condensing a polymer with an oppositely charged polymer to form a particle and sequentially adding oppositely charged polymers to the particle forming at least three layers of polymers. The process is used to form a composition for delivering a biologically active compound to a cell.

Abstract: Administration of expressible polynucleolides encoding eukaryotic heat shock proteins to mammalian cells leads to the stimulation of an immune response to antigens present in those cells. This makes it possible to stimulate an immune response to target antigens, including target tumor antigens or antigens associated with an infectious disease, without having to isolate a unique antigen or antigen-associated heat shock protein for each target antigen by administering to a mammalian subject or to a group of mammalian cells containing the antigen, an expressible polynucleotide encoding a heat shock protein. The expressed heat shock protein may have the same structure as native heat shock proteins, or may have a modified form adapted to control the trafficking of the expressed heat shock protein within the cells.

Abstract: A method of transferring a gene to vertebrate cells is disclosed. The method comprises the steps of: (a) providing microprojectiles, the microprojectiles carrying polynucleic acid sequences, the sequences comprising, in the 5′ to 3′ direction, a regulatory sequence operable in the tissue cells and a gene positioned downstream of the regulatory sequence and under the transcriptional control thereof; and (b) accelerating the microprojectiles at the cells, with the microprojectiles contacting the cells at a speed sufficient to penetrate the cells and deposit the polynucleic acid sequences therein. Preferably, the target cells reside in situ in the animal subject when they are transformed. Preferred target cells are dermis or hypodermis cells, and preferred genes for insertion into the target cells are genes which code for proteins or peptides which produce a physiological response in the animal subject.

Abstract: A method of transferring a gene to vertebrate cells is disclosed. The method comprises the steps of: (a) providing microprojectiles, the microprojectiles carrying polynucleic acid sequences, the sequences comprising, in the 5′ to 3′ direction, a regulatory sequence operable in the tissue cells and a gene positioned downstream of the regulatory sequence and under the transcriptional control thereof; and (b) accelerating the microprojectiles at the cells, with the microprojectiles contacting the cells at a speed sufficient to penetrate the cells and deposit the polynucleic acid sequences therein. Preferably, the target cells reside in situ in the animal subject when they are transformed. Preferred target cells are dermis or hypodermis cells, and preferred genes for insertion into the target cells are genes which code for proteins or peptides which produce a physiological response in the animal subject.

Abstract: The present invention provides a method of targeting transient gene expression and stable gene expression from the exogenous administration of a DNA sequence, which sequence is less than a complete genome, wherein said DNA sequence encodes RNA and protein, or RNA only, to differentiate tissue of living organisms wherein said DNA sequence through a jet injector technique, and said DNA sequence of less than a complete genome is expressed in a living organism. The present invention further provides a flexible multi-nozzle injector device with a wide surface area to allow molding of the injector nozzle to the surface contours of the tissue. Another aspect of the present invention provides an injection device having a long nozzle for injection of DNA deep into the host tissue. Also, in a further aspect the present invention provides an injector device modified to be used with and/or inject through an endoscopic device.

Abstract: The activity of an expression product of a gene introduced into a living body can be regulated by the coexistence of a protein (interfering substance) that interferes with the activity of the expression product. Receptors of the expression product and such may be used as the interfering substance. The activity of an overexpressed product of a gene introduced in gene therapy can be regulated. The administration of an interfering substance can be achieved by in vivo expression of the gene encoding the substance.

Abstract: We describe a process for generating multilayer particles comprising condensing a polymer with an oppositely charged polymer to form a particle and sequentially adding oppositely charged polymers to the particle forming at least three layers of polymers. The process is used to form a composition for delivering a biologically active compound to a cell.

Abstract: Certain embodiments of the invention relate to the use of small particles in biological systems, including the delivery of biologically active agents. Some embodiments involve using a collection of particles comprising an agent, a surfactant molecule having an HLB value of less than about 6.0 units, and a polymer soluble in aqueous solution, wherein the collection of particles has an average diameter of less than about 200 nanometers, wherein the agent is a protein, carbohydrate, polypeptide, adjuvant, nucleic acid encoding a protein, visualization agent, and/or a marker.

Abstract: One form of the present invention is directed to a method of remyelinating demyelinated axons by treating the demyelinated axons with oligodendrocyte progenitor cells under conditions which permit remyelination of the axons. Another aspect of the present invention relates to a method of treating a subject having a condition mediated by a loss of myelin or a loss of oligodendrocytes by administering to the subject oligodendrocyte progenitor cells under conditions effective to treat the condition mediated by a loss of myelin or a loss of oligodendrocytes. A further aspect of the present invention relates to an in vitro method of identifying and separating oligodendrocyte progenitor cells from a mixed population containing other mammalian brain or spinal cord cell types. This further aspect of the present invention involves removing neurons and neuronal progenitor cells from the mixed population to produce a treated mixed population.

Abstract: The present invention relates to a colloidal dispersion of amine-terminated silica particles having a narrowly controlled size range in an aqueous phase for use in diagnostic imaging, drug delivery and gene therapy, as well as methods for preparing surface-modified silica particles suitable for use in an aqueous colloidal carrier medium, for preparing a diagnostic or therapeutic agent for targeted delivery to specific anatomical structures of a patient, and for performing a diagnostic or therapeutic procedure by administration to a patient of at least one diagnostic or therapeutic agent coupled with a colloidal dispersion.

Abstract: Disclosed herein is a novel cell line of human pancreatic cells that secrete insulin in a glucose-dependent manner. The cell line comprises pancreatic cells, such as PANC-1 cells, which are transfected so as to express IDX-1 and cultured in GLP-1. The cell line may be used to investigate the function and development of pancreatic cells, as well as to test the efficacy of drugs that stimulate insulin secretion.

Abstract: A method for activating an equine oocyte comprising exposing oocyte to a medium containing a concentration of calcium of at least about 4 mM. Preferably the oocyte is exposed to this concentration of calcium during activation.

Abstract: The present invention relates to polynucleotides encoding immunogenic HIV polypeptides. Uses of the polynucleotides in applications including immunization, generation of packaging cell lines, and production of HIV polypeptides are also described. Polynucleotides encoding antigenic HIV polypeptides are described, as are uses of these polynucleotides and polypeptide products therefrom, including formulations of immunogenic compositions and uses thereof.

Abstract: A method of delivery of reactive substances that are attached to magnetizable needle-like particles using a magneto-mechanical delivery device. The subject method and device can be utilized for the delivery of reactive or other substances, such as DNA via the penetration of a target body. Such penetration of a target or multiple targets can initiate the interaction between the material contained within the target site and the chemical substances delivered by the particles into the targets. In a preferred embodiment, the subject device is portable and does not require electrical power.

Abstract: A ballistic method of introducing nucleic acid into a nematode worm is described which involves bombarding the nematode with a plurality of microprojectiles. Nematode worms transformed according to the method of the invention are also provided.

Abstract: The invention relates to a particulate product comprising at least one microprojectile; characterised in that the or at least one of the microprojectiles comprises silicon. The invention also relates to devices and components used in the microprojectile implantation of the particulate product to a target of cells or target tissue.

Abstract: The invention concerns compositions comprising at least a nucleic acid and a mineral particle having an interchangeable foliate structure, the method for preparing them and their uses for in vivo, in vitro and/or ex vivo nucleic acid transfection.

Abstract: A gene which is used for transfection into a bony tissue with a gene gun. The gene can well repair a cartilage defective portion to a nearly normal state by transfecting it in the bony tissue for transplantation in the cartilage defective portion without affecting the bony tissue.

Abstract: 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, generating thereby a jet by a sudden expansion of metal gas and thereby generating a shockwave on a surface of an opposite side of the metal foil.

Abstract: In this application is described a poxvirus naked DNA vaccine which protects animals against poxvirus challenge comprising IMV and EEV nucleic acids from poxvirus. Methods of use of the vaccine and its advantages are described.

Abstract: The present invention provides a method of treating cancer in a mammal comprising delivering by electroporation an immunocytokine or cytokine gene in an expression plasmid into cells of the mammal.

Abstract: The present invention provides for lipid:nucleic acid complexes that have increased shelf life and high transfection activity in vivo following intravenous injection, and methods of preparing such complexes. The methods generally involve contacting a nucleic acid with an organic polycation to produce a condensed nucleic acid, and then combining the condensed nucleic acid with a lipid comprising an amphiphilic cationic lipid to produce the lipid:nucleic acid complex. This complex can be further stabilized by the addition of a hydrophilic polymer attached to hydrophobic side chains. The complex can also be made specific for specific cells, by incorporating a targeting moiety such as an Fab′ fragment attached to a hydrophilic polymer.

Abstract: The present invention provides a chimeric protein which comprises a mutated DNA methyltransferase portion and a DNA binding protein portion that binds sufficiently close to a promoter sequence of a target gene which promoter sequence contains a methylation site, to specifically methylate the site and inhibit activity of the promoter and thus inhibit expression of the target gene. This invention also provides for a method for inhibiting the expression of a target gene which includes contacting a promoter of the target gene with the chimeric protein, so as to specifically methylate the promoter sequence of the target gene thus inhibiting expression of the target gene.

Abstract: The present invention involves the role of p53 in the differentiation of embryonic tissues. More particularly, the present invention provides methods of the blocking of p53 function in embryonic tissues, and the use of these tissues as screening tools for substances that are capable of overcoming the p53-related block in differentation, both in vitro and in vivo. The similarities between undifferentiated embryonic cells and tumor cells is evident, and thus these assays serve as a model for possible cancer therapeutics. In addition, methods for identifying additional cellular components that interact p53 or p53-related pathways are provided.

Abstract: An electroporation apparatus for introducing exogenous material into cells is described herein. The apparatus comprises first a base member configured for holding a cell support, the cell support having a top surface portion, with the top surface portion configured for carrying adherent cells. The apparatus further comprises an electrode carrier operably associated with the base member, the electrode carrier having a bottom surface portion, a first electrode connected to the electrode carrier, and a second electrode also connected to the electrode carrier. The electrode carrier has a channel formed therein, with the channel positioned between the first electrode and the second electrode, so that exogenous material may be introduced through the channel and into contact with the cells. Methods for introducing exogenous compounds into a cell and for visually detecting the location of binding events within a cell are also disclosed.

Abstract: The inventive vector specifically directs entry into a cell of monocytic origin. The vector is composed of a nucleic acid component, a lysosome evading component and a particle that can be phagocytized. The vector itself, or cells pretreated with the vector, are useful in all gene medicine applications. Because it is specific for monocytic cells, the inventive vector is particularly suited to vaccine applications. Due to the ability of monocytic cells to target tumors, the inventive vector also is suitable for use in anti-tumor applications, including conventional gene therapy.

Abstract: A method of delivery of reactive substances that are attached to magnetizable needle-like particles using a magneto-mechanical delivery device. The subject method and device can be utilized for the delivery of reactive or other substances, such as DNA via the penetration of a target body. Such penetration of a target or multiple targets can initiate the interaction between the material contained within the target site and the chemical substances delivered by the particles into the targets. In a preferred embodiment, the subject device is portable and does not require electrical power.

Abstract: The present invention provides a method of targeting transient gene expression and stable gene expression from the exogenous administration of a DNA sequence, which sequence is less than a complete genome, wherein said DNA sequence encodes RNA and protein, or RNA only, to differentiate tissue of living organisms wherein said DNA sequence through a jet injector technique, and said DNA sequence of less than a complete genome is expressed in a living organism. The present invention further provides a flexible multi-nozzle injector device with a wide surface area to allow molding of the injector nozzle to the surface contours of the tissue. Another aspect of the present invention provides an injection device having a long nozzle for injection of DNA deep into the host tissue. Also, in a further aspect the present invention provides an injector device modified to be used with and/or inject through an endoscopic device.

Abstract: The present invention provides methods for inhibiting proliferation of astrocytes and astrocytic tumor cells. The present invention further provides methods for treating a condition associated with a defect in astrocyte proliferation in a subject, and methods for treating a condition associated with astrocytic tumor cell proliferation in a subject. Additionally, the present invention is directed to pharmaceutical compositions comprising CD81 protein or nucleic acid and a pharmaceutically-acceptable carrier. Finally, the present invention provides a method for determining whether a subject has an astrocytoma, and a method for assessing the efficacy of astrocytoma therapy in a subject.

Abstract: A gene gun is described wherein the contour design of the spray nozzle of the gene gun modifies the operation of the gene gun. A low pressure gas is used to accelerate the micro-particles that are coated with nucleic acid of a foreign gene into cytoplasma or nuclei of an animal or plant cell to express the special protein and to generate the new biological function.

Abstract: A novel method for introduction of an exogenous genetic substance or a physiologically active compound into cells is provided according to this invention. This method can realize introduction of an exogenous genetic substance or a physiologically active compound of large size with a large amount. Such substance is immobilized to beads of sphere fine particles having a particle size of 0.01 &mgr;m to 10 &mgr;m, and bio-beads thus produced are introduced into cells. Bio-beads comprising calcium alginate are particularly useful for the purpose of the present invention.

Abstract: A method is disclosed for the convenient transformation of the somatic cells of animals. Somatic cell transformation is useful for medical and veterinary care of genetic diseases, and other therapeutic or animal improvement purposes. The method makes use of an electric discharge particle acceleration apparatus which can inject very small particles of gold or other dense material carrying genetic constructs coated on them into the living cells of animals. The animals not only live, but there is no visible bruising or bleeding at the site of the treatment. The method is particularly adaptable since the force of the particle injection in such a spark discharge apparatus is adjustable by adjustments to the voltage of the spark discharge.

Abstract: The present invention relates to coated particles comprising a coating and a core particle comprising an active, wherein the coating comprises a gas phase component. The invention also relates to processes for the manufacture of such coated particles comprising (a) providing a coating material comprising a gas phase component and applying the gas containing coating material to a core particle or (b) providing a coating material comprising a gas generating component, applying the coating material to a core particle and treating the coated particles so as to generate a gas from the gas generating component. Furthermore, it also relates to the use of such coated particles in a number of applications.

Abstract: The present invention relates to a process for uniform and efficient delivery of metal projectiles into living cells and tissue of plants and animals, by bombarding the cells with biological coated on metal bead particles. The metal bead particles are heated in dry oven to high temperature, prior to coating with DNA and by substituting ethanol with isopropanol while coating the particles with DNA, to give several fold higher expression of the nucleic acid delivered into cells.

Abstract: The present invention is directed to a method of separating cells of interest which method comprises: determining cells of interest; selecting a promoter specific for the cells of interest; introducing a nucleic acid molecule encoding green fluorescent protein under control of the promoter into a plurality of cells; and separating cells of the plurality of cells that are expressing said green fluorescent protein, wherein the separated cells are the cells of interest.

Abstract: Disclosed are methods for the isolation of primordial germ cells, culturing these cells to produce primordial germ cell-derived cell lines, methods for transforming both the primordial germ cells and the cultured cell lines, and using these transformed cells and cell lines to generate transgenic animals. The efficiency at which transgenic animals are generated by the present invention is greatly increased, thereby allowing the use of homologous recombination in producing transgenic non-rodent animal species.

Abstract: Provided is a method for introducing a substance into a cell, which method comprises: (a) contacting the cell with a recognition agent to bind the recognition agent to a recognition site on the surface of the cell; and (b) separating the recognition agent from the cell thereby forming a hole in the surface of the cell. Kits for use in such methods are also provided.

Abstract: A method of transferring a gene to vertebrate cells is disclosed. The method comprises the steps of: (a) providing microprojectiles, the microprojectiles carrying polynucleic acid sequences, the sequences comprising, in the 5′ to 3′ direction, a regulatory sequence operable in the tissue cells and a gene positioned downstream of the regulatory sequence and under the transcriptional control thereof; and (b) accelerating the microprojectiles at the cells, with the microprojectiles contacting the cells at a speed sufficient to penetrate the cells and deposit the polynucleic acid sequences therein. Preferably, the target cells reside in situ in the animal subject when they are transformed. Preferred target cells are dermis or hypodermis cells, and preferred genes for insertion into the target cells are genes which code for proteins or peptides which produce a physiological response in the animal subject.

Abstract: The present invention relates to a method of transforming cells in which an appropriate quantity of nucleic acid fragments is introduced into the cells. The nucleic acid fragments are introduced in the form of a nucleic acid composition comprising a nitrogen-containing silicone, useful for compacting the nucleic acid fragments; the compositions comprise aggregates of nucleic acid fragments and silicones according to the invention.

Abstract: A particulate complex is provided comprising a particulate complex comprising a nucleic acid and a biodegradable cationized polyhydroxylated molecule, wherein the polyhydroxylated molecule has a charge up to approximately 1.0 meq/g.

Abstract: The present invention relates to a method for introducing nucleic acids into cells for e.g. producing transiently transfected or stably transformed animal cells by using a specifically designed nucleic acid/protein complex comprising in operable linkage to an expressible DNA or to an oligonucleotide a VirD2 protein, preferably together with a VirE2 protein.

Abstract: A dry powder composition comprises nucleic acid constructs dispersed within with a hydrophilic excipient material, where the powder particles have an average size in the range from 0.5 &mgr;m to 50 &mgr;m. Nucleic acid constructs may comprise bare nucleic acid molecules, viral vectors, or vesicle structures. The hydrophilic excipient material will be selected to stabilize the nucleic acid molecules in the constructs, enhance dispersion of the nucleic acid in dry powder aerosols, and enhance wetting of the nucleic acid constructs as they are delivered to moist target locations within the body.

Abstract: The present invention relates to transfected primary and secondary somatic cells of vertebrate origin, particularly mammalian origin, transfected with exogenous genetic material (DNA) which encodes a desired (e.g., a therapeutic) product or is itself a desired (e.g., therapeutic) product, methods by which primary and secondary cells are transfected to include exogenous genetic material, methods of producing clonal cell strains or heterogenous cell strains, methods of gene therapy in which the transfected primary or secondary cells are used, and methods of producing antibodies using the transfected primary or secondary cells.

Abstract: Disclosed are methods for the isolation of primordial germ cells, culturing these cells to produce primordial germ cell-derived cell lines, methods for transforming both the primordial germ cells and the cultured cell lines, and using these transformed cells and cell lines to generate transgenic animals. The efficiency at which transgenic animals are generated by the present invention is greatly increased, thereby allowing the use of homologous recombination in producing transgenic non-rodent animal species.

Abstract: The present invention provides biodegradable polymer nanospheres capable of transporting and releasing therapeutic agents, specifically nucleic acids. In preferred embodiments, a sub-150 nm nanosphere is formed containing nucleic acids. Thereafter, the agent is released from the nanosphere. In one embodiment a biodegradable polymer nanosphere surface has attached to it a targeting moiety. In another embodiment, the biodegradable polymer nanosphere surface has attached to it a masking moiety. In yet another embodiment both targeting and masking moieties are attached to the nanosphere surface.

Abstract: An improved particle bombardment device for transporting biological substances such as DNA into living cells. The device has a flexible barrel (40) that facilitates endoscopic particle bombardment of in vivo cells without a significant concomitant blast effect and without a need for a vacuum. The device also involves a unique tapered particle-carrying macroprojectile that can travel through convolutions of such a flexible barrel (40) with minimal friction.