Abstract: A process is provided for selectively removing protein aggregates from a protein solution in a normal flow (NFF) filtration process. Preferably, it relates to a process for selectively removing protein aggregates from a protein solution in a normal flow (NFF) filtration process and virus particles from a protein solution in a two-step filtration process. In a first step, a protein solution is filtered through one or more layers of adsorptive depth filters, charged or surface modified microporous membranes or a small bed of chromatography media in a normal flow filtration mode of operation, to produce a protein aggregate free stream. The aggregate free protein stream can then be filtered through one or more ultrafiltration membranes to retain virus particles at a retention level of at least 3 LRV and to allow passage therethrough of an aggregate free and virus free protein solution.

Abstract: An effective prophylactic mucosal gene expression vaccine (GXV), made up of a cocktail of at least 4 different plasmid DNAs encoding corresponding RSV antigens, coacervated with chitosan to formulate nanospheres. In a murine model of RSV infection, intranasal administration with GXV results in significant induction of RSV-specific antibodies, nasal IgA antibodies, cytotoxic T lymphocytes, and IFN-? production in the lung and splenocytes. A single dose of GXV induces a drastic reduction of viral titers.

Abstract: The invention provides attenuated vaccinia virus vaccines that can be used in methods to prevent or treat small pox in patients, as well as methods of obtaining such vaccines.

Abstract: The present invention relates to a formulation, in particular an aqueous formulation comprising (i) a poxvirus of one of the genera orthopoxvirus, avipoxvirus, parapoxvirus, capripoxvirus and suipoxvirus, (ii) a disaccharide, (iii) a pharmaceutically acceptable polymer and optionally (iv) a buffer. The aqueous formulation is particularly suitable for freeze drying processes resulting in a stable, freeze-dried, poxvirus containing composition. The invention further concerns a method for preparing a freeze-dried, poxvirus containing composition and the thus obtained product.

Abstract: The invention includes methods and compositions for the production of high titer recombinant adeno-associated virus (rAAV). The disclosed rAAV are useful in gene therapy applications. Methods are based on the use of recombinant herpes virus vectors and result in highly efficient production of rAAV.

Abstract: The present invention discloses nucleic acid sequence which encodes infectious hepatitis C virus of strain HC-J6CH, gentotype 2a, and the use of the sequence, and polypeptides encoded by all or part of the sequence, in the development of vaccines and diagnostics for HCV and in the development of screening assays for the identification of antiviral agents for HCV.

Abstract: Provided herein are methods for producing alphavirus replicon particles in high yield; replicon RNAs are electroporated into permissive cells, where the cells are at a relatively high density, together with at least one helper nucleic acid providing the necessary functions for packaging. After a growth period in appropriate medium, alphavirus replicon particles are harvested from the surfaces of the cells in which they were produced using a salt wash in which the salt concentration is from about 0.2 to about 5 M sodium chloride, calcium chloride, magnesium chloride, potassium chloride, ammonium acetate, ammonium bicarbonate, among others. After dilution, if necessary, the particles can be purified by a suitable chromatographic technique.

Abstract: The present invention relates first, to the identification of a novel retrovirus and the novel nucleotide sequences encoding a retroviral polymerase gene (POL nucleotides) associated with the existence of MDS and MDS associated disorders. The present invention further relates to methods for using the MDS associated retroviral nucleotides for the detection of MDS and MDS associated disorders in patient samples. The present invention also relates to methods for using and targeting the MDS associated retroviral POL nucleotides in gene therapy protocols for the treatment of MDS and MDS associated disorders in patients in need of such treatment. The present invention further relates to diagnostic protocols and kits for the detection of MDS and MDS associated disorders in tissue samples.

Abstract: Method for efficiently expressing a protein in an active form by using baculovirus expression system wherein the protein is selected from a membrane-bound enzyme, a substrate of the membrane-bound enzyme, a membrane-bound enzyme activator, a membrane-bound transport protein, a channel protein, a membrane structural protein, a protein involved in adhesion, a protein involved in antigen presentation, or a protein involved in formation of high dimensional structure of a protein. There is provided a method for expressing such a protein by culturing a host infected with at least one type of recombinant baculovirus which contains a gene encoding the protein, wherein the protein is expressed in a budded baculovirus released from the host.

Abstract: The present invention encompasses methods of preparing a recombinant viral vector, within a procaryotic cell, by intermolecular homologous recombination, methods of preparing an infectious viral particle containing the recombinant viral vector and pharmaceutical compositions comprising said vector or particle. The invention also encompasses the therapeutic use of said vector or particle, especially in human gene therapy.

Abstract: The present invention relates to a method for the recovery of poxviruses, in particular modified Vaccinia virus Ankara (MVA), from infected cells. According to the present invention the virus-infected cells are subjected to a high-pressure homogenization to obtain a virus containing homogenate. The virus containing homogenate can be subjected to at least one purification step to obtain apoxvirus-enriched fraction. The invention further relates to the virus containing fraction and the virus containing homogenate obtained by the method according to the present invention.

Abstract: The present invention relates to the general field of recombinant protein expression, purification of recombinant proteins, diagnosis of HCV infection, prophylactic treatment against HCV infection and to the prognosing/monitoring of the clinical efficiency of treatment of an individual with chronic hepatitis, or the prognosing/monitoring of the natural disease. In particular, the present invention relates to the use of yeast, i.e. Hansenula or Saccharomyces glycosylation minus strains, for the efficient expression of HCV envelope proteins that are core-glycosylated, purification methods for these proteins, and the use in various applications, such as the use in diagnosis, prophylaxis or therapy of HCV envelope proteins purified according to the present invention.

Abstract: The present invention relates to mutant MVA vaccinia viruses, which are used for the generation of recombinant MVA viruses, as well as host cells, which have been infected with these mutant MVA viruses. The present invention further relates to DNA-vector constructs, and a method for the generation of recombinant MVA by using the mutant MVA viruses and the DNA-vector constructs. The mutant MVA vaccinia viruses of the present invention are characterized in that the MVA ORF 050L gene or a functional part thereof has been inactivated in the viral genome.

Abstract: HCV variants are described. The variants include polynucleotides comprising non-naturally occurring HCV sequences and HCV variants that have a transfection efficiency and ability to survive subpassage greater than HCV that have wild-type polyprotein coding regions. Expression vectors comprising the above polynucleotides and HCV variants are also described, as are the provision of cells and host cells comprising the expression vectors. Methods for identifying a cell line that is permissive for infection with HCV are also provided, as are vaccines comprising the above polynucleotides in a pharmaceutically acceptable carrier. Additionally, methods for inducing immunoprotection to HCV in a primate are described, as are methods for testing a compound for inhibiting HCV replication.

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: The present invention provides compositions useful in and methods for producing populations of infectious, replication-defective alphavirus replicon particles that contain no replication-competent alphavirus particles, as determined by passage on cells in culture. The compositions include helper and replicon nucleic acid molecules that can further reduce the predicted frequency for formation of replication-competent virus and can optimize manufacturing strategies and costs.

Abstract: Described herein is an invention that relates to chemiluminescence-directed antiviral activities of natural and synthesized light-sensitive compounds. Methods are described herein for inactivating infectious virus particles outside and inside an organism. These methods incorporate coupling the antiviral activity of various light-sensitive compounds with chemiluminescence directed by native as well as foreign enzymes of the organism and enhanced by the addition of various anti-quenchers and wavelength-shifting compounds. The methods described herein feature light-sensitive compounds with known antiviral activity exhibited in the presence of light.

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: Described are simplified and efficient methods for preparing recombinant adenovirus using liposome-mediated cotransfection and the direct observation of a cytopathic effect (CPE) in the transfected cells. Also disclosed are compositions and methods involving novel p53 adenovirus constructs, including methods for restoring p53 function and tumor suppression in cells and animals having abnormal p53.

Abstract: Presented herein is a description for the manufacturing of inactivated HIV for use in vaccines against AIDS, as well as other inactivated viruses for other infectious diseases. This invention incorporates methods for inactivating infectious virus particles while retaining protein integrity and antigenicity. The methods utilize critical, near-critical or supercritical fluids with or without polar cosolvents. This invention would allow for the creation of HIV vaccines from genetically attenuated HIV strains for a greater degree of product safety, and from combinations of different HIV strains for broader protection. This HIV vaccine manufacturing technology is inexpensive, amenable to large-scale processing and portable, i.e. it can be readily implemented in a host country site. This invention can be utilized for other viral and bacterial infectious diseases, such as influenza and hepatitis.

Abstract: Immunogenic preparations and vaccines, in particular which are inactivated, effective against feline calicivirosis, based on an FCV virus strain 431 as deposited at the CNCM under the accession number CNCM I-2166, or one of its equivalents, in a veterinarily acceptable vehicle or excipient, preferably combined with FCV virus obtained from another FCV strain, in particular strain G1 as deposited at the CNCM under the accession number CNCM I-2167.

Abstract: The present invention relates to a method of inactivating enveloped viruses in a viral preparation predominantly containing non-enveloped viruses by the action of a solvent at a temperature of between ?5° C. and +50° C. and at a pH of between about 5 and 9. Its subject is also a method of preparing a viral preparation comprising such a method of inactivation. The invention also relates to a viral preparation obtained according to the method of the invention. Finally, it relates to a host cell and a composition comprising such a viral preparation as well as their uses for therapeutic or prophylactic purposes.

Abstract: The present invention relates to the purification of large scale quantities of active (infectious) adenovirus and AAV, especially for use in therapeutic applications. In particular, the invention provides improved methods for contacting such viruses with suitable chromatographic materials in a fashion such that any damage to the virus, particularly to surface components thereof, resulting from contact with such chromatographic materials is minimized or eliminated. The result is the ability to rapidly and efficiently purify commercial level quantities of active (infectious) virus suitable for use in therapeutic applications, e.g. gene transfer/therapy procedures.

Abstract: This invention relates to a method for producing virus RNA polymerases of RNA viruses, more specifically, virus RNA polymerases of RNA viruses free of virus genomic RNA. The methods described in this invention includes the procedures for preparation of cDNAs for the genes for the component proteins of RNA polymerase of an RNA virus, incorporation of the cDNA into baculovirus genome to construct recombinant virus, and the infection of insect cells with the recombinant virus to express RNA polymerase. In this method, it is recommended that all species of the recombinant viruses, each of which is designed for expressing each of the above-mentioned component protein genes of RNA polymerase, are coinfected into insect cells. Thus, cDNA is prepared for each of the component proteins of RNA polymerase and incorporated into baculovirus genome to construct recombinant virus for independently expressing the corresponding protein. In addition, the RNA viruses described above include influenza virus especially.

Abstract: The present invention relates to molecular approaches to the production of nucleic acid sequences which comprise the genomes of chimeric hepatitis C virus-bovine viral diarrhea viruses (HCV/BVDV). The invention also relates to the use of these chimeric nucleic acid sequences to produce chimeric virions in cells and the use of these chimeric virions in HCV antibody neutralization assays, and for the development of vaccines and therapeutics for HCV.

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 provides cell lines which are useful for the rapid detection and production of influenza and parainfluenza viruses. In particular, the invention relates to transgenic mink lung cells which show increased sensitivity to infection by influenza A, influenza B, or parainfluenza 3 viruses, or which are capable of enhanced productivity of infectious virions. The invention is suitable for use in culturing clinical influenza and parainfluenza virus isolates and for the production of influenza and parainfluenza virus for vaccine formulations, as antigen preparations for diagnostic applications, and for screening antiviral drugs.

Abstract: Novel magnetic assay methods and systems. According to a preferred embodiment, a chromatographic medium, which preferably comprises a test strip, is provided that is designed to be contacted with a test solution having activated magnetic particles such that the solution flows bilaterally thereacross. A magnetic field, generated by a magnet or electromagnet, is selectively applied to the medium which causes the charged particles to become substantially bound at a site on the medium specified by the position of the magnet, to thus form a captured line or zone. In one preferred embodiment, the magnetic field is applied at the site on the medium at which the test solution is contacted. The degree of magnetic force applied to the membrane may be selectively adjusted to vary the width or surface area of the capture line or zone.

Abstract: A process for the production of inactivated Hepatitis A virus substantially free of host cell contamination is described, the process comprising: a) culturing Hepatitis A virus and harvesting a Hepatitis A preparation; b) treating said Hepatitis A preparation with a protease; and thereafter c) separating intact virus from protease digested material; d) inactivating said virus. Also described are vaccines comprising the inactivated Hepatitis A virus, preferably in combination with strong adjuvants.

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: Method for preparing a factor VIII solution that is essentially free of viruses and essentially devoid of vWF (von Willebrand factor) and factor VIII-vWF complexes by (a) obtaining a starting factor VIII solution devoid of factor VIII-vWF complexes; and (b) filtering the solution through a hydrophilic virus filter.

Abstract: The present invention provides improved methods of production of viral antigen on a culture of adherent cells bound to a microcarrier, wherein the methods provide for increased viral antigen yield per culture medium volume. The invention is also directed to a cell culture biomass of adherent cells having increased cell density and microcarrier concentration compared to the respective confluent cell culture.

Abstract: Large-scale packaging of alphavirus replicons is accomplished by co-transfecting host cells with three RNA molecules: (1) an alphavirus replicon comprising a sequence encoding a heterologous protein; (2) a helper RNA encoding a capsid protein and (3) a helper RNA encoding two alphavirus glycoproteins. The helper RNAs contain cis-acting elements that allow efficient replication of the helper RNAs and their packaging into viral particles as well as packaging of replicon genomes. These populations of viral particles can be propagated at high titers in cell culture by infecting cells at high multiplicity. Propagation of packaged replicons at an escalating scale is useful for large-scale production of recombinant proteins and/or vaccine.

Abstract: Methods for efficient production of recombinant AAV employ a host cell containing a first nucleic acid molecule comprising from 5? to 3?, a parvovirus P5 promoter, a spacer, an AAV rep sequence and an AAV cap gene sequence, wherein said spacer is of sufficient size to reduce expression of the rep78 and rep68 gene products; a second nucleic acid molecule comprising a minigene comprising a transgene flanked by AAV inverse terminal repeats (ITRs) and under the control of regulatory sequences directing expression thereof in a host cell; and helper functions essential to the replication and packaging of rAAV, which functions are not provided by the first or second nucleic acid molecules. Host cells and molecule constructs are also described.

Abstract: The present invention relates to a process for producing poxvirus, in particular Chordopoxvirus, wherein the poxvirus is cultivated at a temperature below 37° C. The process leads to increased virus propagation at the decreased temperature.

Abstract: The invention concerns a method for producing recombinant adenovirus by which viral DNA is introduced in a packaging cell culture and the viruses produced are harvested after liberation in the supernatant. The invention also concerns the viruses produced and their use.

Abstract: A flexible automated apparatus for isolating and purifying viruses, proteins and peptides of interest from a plant material is disclosed, the apparatus being applicable for large scale purification and isolation of such substances from plant material. The flexible automated apparatus provides an efficient apparatus for isolating viruses, proteins and peptides of interest with little waste material. The automated apparatus for isolating viruses, proteins and peptides of interest includes a grinding apparatus for homogenizing a plant to produce a green juice, a means for adjusting the pH of and heating the green juice, a means for separating the target species, either virus or protein/peptide, from other components of the green juice by one or more cycles of centrifugation, resuspension, and ultrafiltration, and finally purifying virus particles by such procedure as PEG-precipitation or purifying proteins and peptides by such procedures as chromatography and/or salt precipitation.

Abstract: A method for classifying and counting leukocytes comprises the steps of: (1) adding to a hematological sample the following fluorescence-labeled antibodies labeled with fluorescent dyes which emit fluorescences distinguishable from each other; (a) a first fluorescence-labeled antibody (1st antibody) which bonds specifically to leukocytes, (b) a second fluorescence-labeled antibody (2nd antibody) which bonds to at least one kind of neutrophilic cells, and (c) a third fluorescence-labeled antibody (3rd antibody) which bonds to at least one kind of immature granulocytic cells, in order to stain leukocytic cells in the sample, and removing erythrocytes from the sample; (2) analyzing the resulting sample using a flow cytometer to measure at least one scattered light signal and three separate fluorescence signals; (3) defining a group of granulocytic cells on the basis of intensity of the scattered light and intensity of fluorescence from the 1st antibody; (4) defining neutrophilic cells in the defined group of gra

Abstract: A new mumps vaccine is presented, comprising a homogeneous pure isolate derived from the Jeryl-Lynn strain of mumps virus. In a preferred embodiment of the invention the vaccine produces higher seroconversion and anitbody titres than know commercial vaccines.

Abstract: Recombinant negative-strand viral RNA templates are described which may be used with purified RNA-directed RNA polymerase complex to express heterologous gene products in appropriate host cells and/or to rescue the heterologous gene in virus particles. The RNA templates are prepared by transcription of appropriate DNA sequences with a DNA-directed RNA polymerase. The resulting RNA templates are of the negative-polarity and contain appropriate terminal sequences which enable the viral RNA-synthesizing apparatus to recognize the template. Bicistronic mRNAs can be constructed to permit internal initiation of translation of viral sequences and allow for the expression of foreign protein coding sequences from the regular terminal initiation site, or vice versa.

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 invention relates to assembled particles of a plant virus containing a foreign peptide insert in the coat protein of the virus. The site of the insert is preferably free from direct sequence repeats flanking the insert. The invention relates to a method of production of the particles and their use, in particular in vaccines.

Abstract: The invention involves a papilloma pseudo-virus that can induce immune response after oral intake as well as its preparation. It is characterized in that HPV or BPV pseudo-virus are made by disrupting HPV-VLP or BPV-VLP, mixing them with plasmids (plasmids or DNA vaccine), and reassembling them into the pseudo-viruses (VLPs with plasmids inside). Oral administration of the pseudo-viruses will result in delivery to mucosal and systemic lymphoid tissues and induce immune responses for disease prevention and treatment. The pseudo-virus induces stronger immune response than DNA vaccines. Additionally, the pseudo-virus can be applied in gene therapy by bringing the therapeutic genes into lymphoid tissues in the human body.

Abstract: A method to prepare viruses lacking ion channel activity is provided.

Abstract: The present invention relates, to novel methods and substrates for the propagation of viruses. The invention relates to IFN-deficient substrates and methods for propagating viruses in these unconventional substrates. In particular, the invention relates to methods of propagating viruses in immature embryonated eggs, preferably six- to nine-day-old chicken eggs. The methods of the invention are particularly attractive for growing viruses suitable for use in vaccine and pharmaceutical formulations.

Abstract: The present invention provides a novel inactivated virus particle and a reinforced immunogen which have a reinforced titer about twice to about 10 times that of a conventional vaccine, as well as a method for producing the same. The inactivated virus particle is produced by inactivating and then purifying a culture by physical means. The inactivated virus particle of the present invention is useful in a diagnostic agent for infectious disease caused by a group of Japanese encephalitis virus.

Abstract: The invention relates to a process for separating viruses of different sizes, with the virus-containing solution preferably being filtered through one or more filter membranes.

Abstract: Sample pretreatment solutions for influenza virus tests by immunochromatography are described.

Abstract: The present invention relates to a method for the recovery of poxviruses, in particular modified Vaccinia virus Ankara (MVA), from infected cells. According to the present invention the virus-infected cells are subjected to a high-pressure homogenization to obtain a virus containing homogenate. The virus containing homogenate can be subjected to at least one purification step to obtain apoxvirus-enriched fraction. The invention further relates to the virus containing fraction and the virus containing homogenate obtained by the method according to the present invention.

Abstract: Methods and compositions for the optimization of production of influenza viruses suitable as influenza vaccines are provided.