Abstract: This invention provides methods and compositions for producing high titer, substantially purified preparations of recombinant adeno-associated virus (AAV) that can be used as vectors for gene delivery. At the onset of vector production, AAV producer cells of this invention typically comprise one or more AAV packaging genes, an AAV vector comprising a heterologous (i.e. non-AAV) transgene of interest, and a helper virus such as an adenovirus. The AAV vector preparations produced are generally replication incompetent but are capable of mediating delivery of a transgene of interest (such as a therapeutic gene) to any of a wide variety of tissues and cells. The AAV vector preparations produced according to this invention are also substantially free of helper virus as well as helper viral and cellular proteins and other contaminants. The invention described herein provides methods of producing rAAV particles by culturing producer cells under conditions, such as temperature and pH, that promote release of virus.

Abstract: Disclosed are nucleic acid molecules encoding novel DKR polypeptides. Also disclosed are methods of preparing the nucleic acid molecules and polypeptides, and methods of using these molecules.

Abstract: A process for the inactivation and/or elimination of coated and/or noncoated viruses from a plasma protein solution by addition of an ammonium salt at alkaline pHs is described, in which a chromatographically prepurified plasma protein solution is subjected to an incubation at room temperature and at an ammonium salt concentration of 13 to 22% by weight, pasteurized at approximately 60° C. for several hours after separation of the precipitates and removal of the ammonium salt down to a residual content of less than 0.5 mol/l and then processed to give a therapeutically employable plasma protein preparation.

Abstract: A method for increasing production yield of viruses, viral proteins, and other related biological materials through enhanced control and stabilization of protein production via stress proteins and the resultant protein products. The present invention is also directed to methods for selection or engineering of cell lines yielding such enhanced stabilized products. More specifically, example embodiments of the present invention are directed to methods for enhancing production of a viral agent, production of cell lines exhibiting permanent genetic modification, production of permissive eucaryotic cell lines, enhancing functional recombinant product yield, and the products of such methods.

Abstract: Polynucleotide encoding modified HIV Env polypeptides are disclosed. The Env polypeptides are modified so as to expose at least part of the CD4 binding region. Methods of diagnosis, treatment and prevention using the polynucleotides and polypeptides are also provided.

Abstract: The present invention is directed to novel bacteriophage compositions useful in treating food products to prevent bacterial contamination.

Abstract: This invention relates to a method for culturing a virus including the steps of: (A) providing cells from a cell line susceptible to infection by the virus and a specimen; (B) treating the cells with a compound of formula RC(O)Q, wherein Q is R, OR, OX or X, each R is independently hydrogen or a hydrocarbyl group containing 1 to about 10 carbon atoms and wherein X is hydrogen or a cation; © inoculating the treated cells with the specimen; and (D) incubating the inoculated cells to allow viral growth to proceed.

Abstract: The invention provides a method for producing purified replication-defective recombinant AAV virions. The method comprises introducing into a suitable host cell an AAV vector, an AAV helper construct and an adenoplasmid accessory construct into the host cell. The adenoplasmid accessory plasmid is composed adenovirus plasmid DNA unable to be packaged into adenoviral particles because it lacks packaging signal sequence(s) or it contains additional sequences making it too large to package. The host cell is cultured to produce crude rAAV virions and then lysed. The resulting cell lysate is applied to a chromatographic column containing sulfonated cellulose or subjected to cesium chloride equilibrium gradient centrifugation and the purified rAAV virions are recovered.

Abstract: This invention provides methods and compositions for producing high titer, substantially purified preparations of recombinant adeno-associated virus (AAV) that can be used as vectors for gene delivery. At the onset of vector production, AAV producer cells of this invention typically comprise one or more AAV packaging genes, an AAV vector comprising a heterologous (i.e. non-AAV) transgene of interest, and a helper virus such as an adenovirus. The AAV vector preparations produced are generally replication incompetent but are capable of mediating delivery of a transgene of interest (such as a therapeutic gene) to any of a wide variety of tissues and cells. The AAV vector preparations produced according to this invention are also substantially free of helper virus as well as helper viral and cellular proteins and other contaminants. The invention described herein provides methods of producing rAAV particles by culturing producer cells under conditions, such as temperature and pH, that promote release of virus.

Abstract: The invention relates to a process for recombinantly preparing picornavirus particles, in particular hepatitis A virus particles, their precursors and particles which are derived therefrom, with a structural protein precursor molecule (P1-2A or P1) and the corresponding P3 region (3ABCD) being coexpressed in cis or in trans.

Abstract: The present invention provides experimentally-generated cold-adapted equine influenza viruses, and reassortant influenza A viruses comprising at least one genome segment of such an equine influenza virus, wherein the equine influenza virus genome segment confers at least one identifying phenotype of the cold-adapted equine influenza virus, such as cold-adaptation, temperature sensitivity, dominant interference, or attenuation. Such viruses are formulated into therapeutic compositions to protect animals from diseases caused by influenza A viruses, and in particular, to protect horses from disease caused by equine influenza virus. The present invention also includes methods to protect animals from diseases caused by influenza A virus utilizing the claimed therapeutic compositions.

Abstract: Human papillomavirus virus-like particles (VLPs) are subjected to various maturation conditions, including incubation at higher temperatures, exposure to soluble metals or thios-oxidation. The resultant matured VLPs are more stable, and can be used to make a vaccine formulation with increased shelf life and higher potency.

Abstract: The present invention describes (1) an immortal cell line derived from grouper and a method for establishing the cell line; (2) methods for mass producing and purifying aquatic viruses using the immortal cell line from grouper; (3) an anti-NNV antibody and a method for producing the anti-NNV antibody; and (4) a vaccine of NNV and a method for protecting fish against NNV infection. The present immortal cell line is derived from the grouper and is susceptible to the viral families of Birnaviridae such as Infectious Pancreatic Necrosis Virus (IPNV); Herpesviridae such as Eel Herpes Virus Formosa (EHVF); Reoviridae such as Hard Clam Reovirus (HCRV); and Nodaviridae such as Nervous Necrosis Virus (NNV).

Abstract: The present invention relates to a method for helper virus-free packaging of gene vector DNA into the virus particles of a DNA virus as well as to eukaryotic helper cells for helper virus-free packaging of gene vector DNA into the virus particles of a DNA helper virus wherein a DNA virus having a genome ≧100 kbp is employed (FIG. 1).

Abstract: Culture of human oranimal herpesviruses, e.g. disabled mutant herpesviruses, can be carried out on primary cells which have been made recombinant so as to express a first gene that extends their culturable life and a second gene derived from the virus. The virus products can be used in vaccines or gene delivery to cells.

Abstract: An infectious clone based on the genome of a wild-type RNA virus is produced by the process of providing a host cell not susceptible to infection by the wild-type RNA virus, providing a recombinant nucleic acid based on the genome of the wild-type RNA virus, transfecting the host cell with the recombinant nucleic acid and selecting for infectious clones. The recombinant nucleic acid comprises at least one full-length DNA copy or in vitro-transcribed RNA copy or a derivative of either. The infectious clones can be used in single or dual purpose vaccines and in viral vector vaccines.

Abstract: The present invention relates to the use of virus-like particles (VLP's) of papillomavirus for preparing vector pseudoviruses useful for transferring genetic material into target cells of an organism

Abstract: A method of reducing levels of E. coli O157 strains within the gastrointestinal tract of a ruminant animal using specific bacteriophage(s) is herein described. Also described is a pharmaceutical composition comprising at least one of said bacteriophages and a method for isolating or selecting bacteriophages useful in reducing E. coli O157 levels as described above.

Abstract: Novel cell lines useful for trans-complementing E1-deleted adenoviral vectors are described. The cell lines are capable of providing high yields of E1-deleted adenoviral vectors in the absence of replication-competent adenovirus over multiple passages.

Abstract: A method of disassembly/reassembly of papillomavirus VLPs is provided. The resultant VLPs have enhanced homogeneity, present conformational, neutralizing PV epitopes, and therefore are useful prophylactic and diagnostic agents. Further, these VLPs can be used to encapsulate desired moieties, e.g., therapeutic or diagnostic agents, or “marker” DNAs, and the resultant VLPs used as in vivo delivery vehicles or as pseudovirions for evaluating vaccine efficacy.

Abstract: The present invention provides compositions and methods of producing recombinant AAV (rAAV) virions in large amounts or high titers. Also provided are methods for producing stably transformed host cells capable of producing rAAV virions.

Abstract: The present invention is directed to attenuated pestivirus mutants, which have a reduced ability to replicate as exhibited by a small plaque size. The mutations are in the 5′ nontranslated region of the viral genome. These mutant viruses are useful as live vaccines in the control of bovine viral diarrhea, border disease and classical swine fever.

Abstract: The invention provides new methods for purifying and concentrating viruses. The inventors have discovered that high molecular weight proteoglycans present in retroviral stocks are co-concentrated with the retroviruses, and can inhibit retroviral transduction. The new purification and concentration methods feature treatment of virus stock with an anionic polyelectrolyte and a cationic polyelectrolyte, followed by centrifugation. The new methods minimize the amount of proteoglycan co-precipitated with the infectious virus.

Abstract: The present invention relates to avian cell lines which efficiently support the growth and productive infection of Marek's Disease Virus at high titers. The present invention also relates to avian cell lines which have been engineered to support the growth and productive infection of recombinant Marek's Disease Virus at high titers. The present invention relates a process for the preparation of Marek's Disease Virus in quantities suitable for vaccine purposes.

Abstract: Primary receptors and co-receptors for adeno-associated virus (AAV) attachment to and infection of target cells are described. Such receptors can be used to facilitate AAV attachment to and infection of cells, e.g., for gene therapy. Methods for purification and/or concentration of AAV are also described. Methods of facilitating or enhancing AAV infection of a cell are also provided. Also described are methods of inhibiting or preventing infection of AAV into a cell. Cell samples may be screened for permissiveness for AAV attachment and infection by detecting the presence or abundance of cellular receptors that mediate attachment and/or infection of AAV into the cell. Formulations and kits for mediating AAV attachment to, and infection of, cells are also provided herein.

Abstract: The present invention provides two new HIV/SIV translocation promoting agents, Bonzo and BOB. The present invention also provides the amino acid and DNA sequences of human, African green monkey, and pigtail macaque of the receptor protein Bonzo. Mammalian cells transfected with Bonzo and/or BOB and human CD4 as well as antibodies to the receptor Bonzo are also included. Furthermore, a method of identifying other such translocation promoting agents is also disclosed. Diagnostic and therapeutic uses of the translocation promoting agents of the present invention are also provided. Furthermore, the present invention provides methods of identifying agents that can modulate the expression and/or function of Bonzo/STRL33. Such agents can be used to either treat inflamation, or alternatively, to enhance the immune response.

Abstract: Crystal growth can be initiated and controlled by dynamically controlled vapor diffusion or temperature change. In one aspect, the present invention uses a precisely controlled vapor diffusion approach to monitor and control protein crystal growth. The system utilizes a humidity sensor and various interfaces under computer control to effect virtually any evaporation rate from a number of different growth solutions simultaneously by means of an evaporative gas flow. A static laser light scattering sensor can be used to detect aggregation events and trigger a change in the evaporation rate for a growth solution. A control/follower configuration can be used to actively monitor one chamber and accurately control replicate chambers relative to the control chamber. In a second aspect, the invention exploits the varying solubility of proteins versus temperature to control the growth of protein crystals.

Abstract: Filtration methods comprise virus-filtering a solution containing at least one macromolecule. The total salt content of the solution is within the range of from about 0.2M to 2M or within the range of from about 0.2M up to saturation with the salt.

Abstract: Methods for purifying adenoviruses from a contaminated sample using a hydroxyapatite medium are provided. Sodium chloride concentrations present in buffers used throughout the method are at least 150 mM to prevent the adenovirus from irreversibly binding to the hydroxyapatite. Levels of contaminants and empty adenovirus capsids from samples purified by conventional purification techniques are further reduced to provide a highly pure adenovirus preparation.

Abstract: Production of enveloped RNA virus-like particles intracellularly in vitro in insect cells using a recombinant baculovirus vector containing a cDNA coding for viral structural proteins is disclosed. In vitro production and purification of hepatitis C virus (HCV)-like particles containing HCV core protein, E1 protein and E2 protein is disclosed. Production of antibodies in vivo to the purified HCV-like particles is disclosed.

Abstract: A method of enriching a solution of an adenovirus comprising applying a mixed solution comprising an adenovirus and at least one undesired type of biomolecule to an anion exchange chromatography resin containing a binding moiety selected from the group consisting of dimethylaminopropyl, dimethylaminobutyl, dimethylaminoisobutyl, and dimethylaminopentyl and eluting the adenovirus from the chromatography resin. Also provided is a method of purifying an adenovirus from adenovirus-infected cells comprising lysing such cells, applying the lysate to a single chromatography resin, eluting the adenovirus from the chromatography resin, and collecting a fraction containing adenovirus that is substantially as pure as triple CsCl density gradient-purified adenovirus.

Abstract: In accordance with the present invention, numerous experiments were conducted to determine the role of infectious bursal disease (IBD) virus in the induction of lesions associated with proventriculitis syndrome in chickens. Parameters examined included age of the chicken at IBD virus exposure, concentration of IBD virus at exposure, the strain of IBD virus, dietary influence in the presence of IBD virus, mixed IBD virus infection, auto-immune mediated IBD reactions associated with lesion production, viral induced apoptotic tissue injury and isolation and characterization of the causative agent. The experiments were carried out in SPF white leghorns, with the experimental birds being examined for the presence of gross and microscopic lesion at 4 and 11 days post challenge. Tissue homogenates were analyzed for the presence of IBDV at 4 and 11 days post challenge with Antigen Capture ELISA (AC-ELISA).

Abstract: A method for detecting mutated alleles in an excess of wild type alleles in a sample by isolating sample DNA; amplifying a target DNA sequence; and separating mutated DNA sequences from wild type DNA sequences by virtue of the preferential binding of the wild type sequences to carrier-bound complementary oligonucleotides. The amplification and separation steps may be iterated through one or more additional cycles to enhance sensitivity.

Abstract: A process is provided for selectively removing protein aggregates and virus particles from a protein solution in a two-step filtration process. In a first step, a protein solution is filtered by tangential flow filtration through a cellulosic ultrafiltrate membrane at a transmembrane pressure of between about 1 and about 10 psi to produce a first permeate and a retentate stream. Water or aqueous buffer is added to the retentate stream to form an essentially constant volume retentate stream. The first permeate is filtered through a second ultrafiltration membrane to retain virus particles at a retention level of at least 3 LRV and to allow passage therethrough of a protein aggregate free and virus free protein solution.

Abstract: Novel cell lines useful for trans-complementing E1-deleted adenoviral vectors are described. The cell lines are capable of providing high yields of E1-deleted adenoviral vectors in the absence of replication-competent adenovirus over multiple passages.

Abstract: A simple and efficient method of producing mammalian reovirus is developed using HEK 293 cells. The method provides for fast production of reovirus in high yield. Furthermore, this method provides for a simpler purification procedure of the produced reovirus.

Abstract: A method of enriching a solution of an adenovirus comprising applying a mixed solution comprising an adenovirus and at least one undesired type of biomolecule to an anion exchange chromatography resin containing a binding moiety selected from the group consisting of dimethylaminopropyl, dimethylaminobutyl, dimethylaminoisobutyl, and dimethylaminopentyl and eluting the adenovirus from the chromatography resin. Also provided is a method of purifying an adenovirus from adenovirus-infected cells comprising lysing such cells, applying the lysate to a single chromatography resin, eluting the adenovirus from the chromatography resin, and collecting a fraction containing adenovirus that is substantially as pure as triple CsCl density gradient-purified adenovirus.

Abstract: Oligonucleotides, for detection of small round structured virus (SRSV) RNA, which bind specifically to SRSV RNA at a relatively low, constant temperature (for example, 41° C.), and an SRSV RNA detection method, which is a constant temperature nucleic acid amplification method employing the oligonucleotides, are provided.

Abstract: An improved species specific vaccine having reduced risks of engendering autoimmune disease in vaccinated animals is described. The vaccine is formulated to be substantially free of heterologous species serum components. Preferably the species specific vaccines are produced from antigen concentrates recovered from cell culture media comprising serum wherein the serum consists essentially of homologous species serum.

Abstract: A new mumps vaccine is presented, comprising a homogeneous pure isolate derived from the Jeryl-Lynn strain of mumps virus.

Abstract: A method for separating microorganisms, especially infectious agents, from a mixture by two dimensional centrifugation on the basis of sedimentation rate and isopycnic banding density, for sedimenting such microorganisms through zones of immobilized reagents to which they are resistant, for detecting banded particles by light scatter or fluorescence using nucleic acid specific dyes, and for recovering the banded particles in very small volumes for characterization by mass spectrometry of viral protein subunits and intact viral particles, and by fluorescence flow cytometric determination of both nucleic acid mass and the masses of fragments produced by restriction enzymes. The method is based on the discovery that individual microorganisms, such as bacterial and viral species, are each physically relatively homogeneous, and are distinguishable in their biophysical properties from other biological particles, and from non-biological particles found in nature.

Abstract: Methods of production and purification for viruses and virus-derived vectors, including those related to alphaviruses, are disclosed. In one aspect, methods of purification that subject alphavirus replicon particle preparations to one or more steps of chromatographic purification, such as using an ion exchange resin, are provided. Also disclosed are methods of characterizing alphavirus replicon particles and utilizing these materials for vaccines and gene-based therapeutics.

Abstract: The invention provides replication of high growth influenza virus strains, derived from clinical isolates, in cultured mammalian cells by infecting the mammalian cells with the high growth strains to obtain infected cells, and culturing the cells while maintaining a trypsin concentration range of 0.05-1.0 &mgr;g/ml in the culture medium, where the resulting replicated virus is suitable for use in mammalian influenza vaccines and vaccination methods, which are also provided by the invention.

Abstract: The present invention relates to live PRRS viruses which are attenuated by amino acid mutations in a specific site of the viral protein coded by the open reading frame (ORF) selected from the group of ORF 1a, ORF 1b and/or ORF 2. The invention also pertains to nucleotide sequences coding said viruses, methods of generating such viruses and their use for the preparation of a pharmaceutical composition for the prophylaxis and treatment of PRRS infections.

Abstract: The present invention relates to methods for the production of high titers of serum-free lytic viruses in a hollow fiber cartridge capillary system. The invention further relates to methods of infecting target cells at high multiplicity and for producing high concentrations of transduced target cells.

Abstract: Improved forms of vaccines which comprise proteosomes and protein antigens are described. Vaccines which contain influenza HA as the antigen are used for illustration as to demonstrate efficacy. Improvements in the preparation of the vaccines themselves and the proteosome component are also included.

Abstract: A protocol for the ion exchange concentration of baculovirus is presented.

Abstract: Methods are disclosed for the purification of encapsulated viruses. The methods are advantageous in that they employ filtration aids, together with low concentrations of metal ions, in place of nucleases for purification. This provides important advantages for commercial scale purification of viruses.

Abstract: A method for determining the number or concentration of virus particles in a sample by use of a light scattering detector. The method may be used to quantitate purified virus preparations or virus samples containing contaminants, including ultraviolet light-absorbing contaminants, such as proteins. The method is useful for quantitation of viruses for use in gene therapy, oncolytic viruses for tumor cell lysis and virus-based vaccines.

Abstract: The invention provides methods for purifying a virus from impurities in an aqueous medium.