112 results about "Virus multiplication" patented technology

The present invention relates to compounds that modulate the replication of negative-sense, single-stranded RNA viruses, such as influenza virus, and the use of such compounds. The invention relates to methods for increasing the titer of negative-sense, single-stranded RNA viruses, such as influenza virus, in substrates for virus propagation (e.g., tissue culture). The invention also relates to the use of compounds that decrease virus replication as antiviral agents. The invention further relates to methods for identifying compounds that modulate the replication of negative-sense, single-stranded RNA viruses, in particular, influenza virus.

The invention provides a method for producing rabies viruses by the suspension culture of BHK21 cells, which comprises a step of performing suspension culture of BHK21 cells in bioreactor containing a BHK21 cell culture medium, wherein the conditions for the suspension culture in the step include a temperature of 32 to 36 DEG C, a pH value of 7.0 to 8.0 and a dissolved oxygen concentration of 30 to 50 percent; and the BHK21 cell culture medium comprises the components shown in a table 1. The method overcomes the biases of the prior art and realizes the production of the rabies viruses by the suspension culture of BHK21 cells in the bioreactor. The obtained rabies viruses can be used for producing rabies vaccine. Due to the automatic culture environment parameter control of the bioreactor, the cells grow and the viruses propagate in more favorable environments, the virus titer is improved and the large-scale automatic continuous production can be realized.

The invention discloses an avian influenza virus inactivated vaccine and a preparation method thereof. Adaptive avian influenza viruses are adaptive to screening and domestication of a continuous cell line, and after the viruses are adaptive to cells, the number of the viruses is continuously increased until the viruses are inoculated into a cell culture bottle or a bioreactor to culture. After the obtained culture is inactivated, the obtained culture and mineral oil adjuvant are mixed and emulsified so as to prepare the vaccine. A virus reproduction method for producing an avian vaccine by means of utilizing a large quantity of chicken embryo is omitted, resource consumption is reduced, environmental pollution is decreased, biosafety is guaranteed, cost is lowered, and production efficiency is improved. By the aid of the preparation method, high-potency viruses can be produced to prepare corresponding vaccines, and requirements of fast and high-efficient vaccine production are met.

The invention discloses a production process of fusion expression recombinant chicken interferon alpha. The process includes the steps of: S1, according to the preference of Escherichia coli codon, conducting codon optimization on a chicken interferon alpha gene sequence published in Genebank, and artificially synthesizing the chicken interferon alpha gene; S2, according to the codon optimized chicken interferon alpha gene, designing three specific primers; S3, constructing recombinant chicken interferon alpha plasmid containing ProS2 dissolution promoting label; S4, transforming and identifying the recombinant expression plasmid; S5, conducting inducible expression of recombinant chicken interferon alpha; S6, extracting an expression product and conducting protein renaturation purification: S61, inclusion body extraction and treatment; S62, inclusion body denaturation; S63, denaturation solution renaturation; and S64, nickel column affinity purification. By means of cell cytopathic inhibition, the invention detects that the interferon has the activity of inhibiting vesicular stomatitis virus proliferation, and the activity unit reaches 7.32*10<7>UI/mg.

The invention relates to the use preferably of at least one active ingredient for the prophylaxis and/or therapy of a viral disease, wherein this active ingredient inhibits at least one component of the cellular signal transduction pathway for the activation of the transcription factor NF-kB such that the virus multiplication is inhibited. The present invention relates furthermore to the local, preferably aerogenic administration of the active ingredient according to the invention for inhibiting a virus multiplication. The active ingredient according to the invention may be combined with at least one further antivirally effective substance for the prophylaxis and/or therapy of a viral disease.

The invention relates to a nasal spray preparation for resisting respiratory virus infection. The preparation comprises liposome-loaded angiotensin converting enzyme 2 (ACE2) recombinant protein, sialic acid, natural polysaccharide and normal saline. The nasal spray preparation is mainly prepared from the liposome-loaded ACE2 extracellular domain recombinant protein, can competitively inhibit thecombination of viruses and host cells, prevents and treats respiratory tract infection caused by various viruses, and enhances the antiviral range and clinical indications of the nasal spray preparation for resisting respiratory tract virus infection. In addition, the nasal spray preparation can be used for directly spraying medicines to virus propagation parts, and is convenient and simple to use.

The invention provides virus-like particles for treatment of viral infections based on the virus causing the infection. The virus-like particles comprise the virus recombinant proteins that form a capsid, recombinant virus membrane proteins attached to the capsid and vRNA packaged within said capsid. The vRNA is generated from a DNA sequence encoding a polypeptide capable of specifically binding to a constant region of a nonstructural protein of the virus that is essential for propagation of the virus.

The invention discloses a production method of a canine parvovirus inactivated vaccine by utilizing a bioreactor. The production method comprises the following steps: (1) culturing a vaccine producing cell by applying a microcarrier and chip carrier system of the bioreactor; (2) vaccinating a canine parvovirus and performing virus multiplication culture; (3) harvesting virus culture fluid; and (4) inactivating the virus fluid with BEI (binary ethyleneimine), and adding adjuvant to prepare the vaccine. The canine parvovirus inactivated vaccine has the advantages of high density of cultured cell, high virus titer, uniform and stable quality, controllable process and high production efficiency, and the defects of large difference among product batches, low antigen content and low production efficiency in a conventional spinner bottle or chick embryo production process can be overcome.

The invention discloses an influenza virus subunit vaccine and a preparation method thereof. The influenza virus subunit vaccine is formed by further purifying cracked viral proteins through a cracking agent and a new purification method; each agent of subunit vaccine comprises more than 80% of H1N1, H3N2 and B type influenza hemagglutinin, and does not comprise adjuvant, thiomersal or other corrosion removers. The invention further provides the preparation method of the influenza virus subunit vaccine. The preparation method of the influenza virus subunit vaccine comprises the following steps of virus inoculation, virus multiplication culture, allantonic fluid harvesting, clarification, inactivation, ultrafiltration and concentration, cracking and overspeed centrifugal purification, gel filtration chromatography purification, mixture, filtration sterilization, subpackage, packaging and the like. The influenza virus subunit vaccine can improve safety, eliminate untoward effects caused by adjuvant and eliminate the toxic and side effects caused by thiomersalate.

The invention provides a method for producing a blue-ear disease vaccine by culturing a sensitive cell. The method comprises the following steps: 1) culturing the sensitive cell on a microcarrier in a bioreactor; 2) inoculating a porcine reproductive and respiratory syndrome virus to the sensitive cell cultured on the microcarrier; 3) continuously culturing the sensitive cell; and 4) obtaining virus solution to produce the blue-ear disease vaccine. The method produces the blue-ear disease vaccine by culturing the sensitive cell in the bioreactor by applying microcarrier culture technology. The microcarrier provides larger surface area, the cell can reproduce to generate higher cell density, and automatically controlled culture environment parameters of the bioreactor ensure that the cell growth and virus propagation can be in an excellent environment, so the method can improve the virus titer, and ensures more stable vaccine quality.

The invention discloses a bombyx mori nuclear polyhydrosis virus (BmNPV) 39k inducible promoter and application thereof. A nucleotide sequence of the promoter is shown as SEQ ID No.1; the promoter has obvious BmNPV inducible promotion activity and can make cells start foreign gene expression when the cells are subjected to the BmNPV infection; an RNAi vector using a BmNPV multiplication essential gene as a target can be built by utilizing the promoter; the abundant expression of shRNA can be performed when the bombyx mori is subjected to the BmNPV infection; the shRNA is cut into siRNA in a cell by an enzyme; specific degradation is performed on the BmNPV multiplication essential gene mRNA to initiate the BmNPV multiplication essential gene to be transcribed and then silenced, so the aim of inhibiting the virus multiplication is fulfilled, and the promoter can be used for preventing and controlling the BmNPV infection of the bombyx mori, performing genetic engineering breeding of bombyx mori anti-BmNPV strains and providing a good reference mode for transgenic therapy of other biologic virus diseases.

The invention provides a method for industrially producing a swine parvovirus vaccine by utilizing a bioreactor, comprising the following steps of: (1) sterilizing a micro-carrier and the bioreactor, adding a cell growth solution, inoculating, preparing the vaccine and culturing with cells, inoculating a swine parvovirus after the cell on the micro-carrier forms a compact single layer, and continuously culturing to propagate the virus; (2) stopping culturing until the cytopathy reaches more than 80%, and harvesting a virus solution; (3) carrying out ultrafiltration concentration and virus inactivation on the harvested virus solution; and (4) purifying and inactivating the virus through a column chromatography method to prepare the vaccine. The invention has the advantages of favorable controllability of processing parameters, high cell density and virus titer, favorable vaccine safety, stable and reliable quality, high production efficiency, and the like.

The invention discloses a method for preparing a chicken infectious bronchitis virus HA antigen. The chicken infectious bronchitis virus HA antigen is prepared by the steps of virus multiplication, virus solution concentration and phospholipase C treatment, and antigen inactivation and stabilization. The stability of the prepared HA antigen at a temperature of 4 DEG C is prolonged to more than 6 months from the original 24 hours; besides, the valence of antigen is stable so as to solve the difficult problems which are not solved for a long time that the antigen is temporarily prepared for the test of each time and needs to be detected before use, and reduce the workload of the HI test. The prepared IBV HA antigen can be successfully applied to detecting the valence of antibody of serum HI after the chicken IBV vaccine immunization, and solves the problem that the detection of the IBV vaccine immunization effect is realized through neutralization tests in China for a long time, thereby wasting time and energy and having the danger of poison diffusion.

The invention belongs to the technical field of preparation of virus vaccine, and concretely discloses an application of an H3N2 canine influenza virus CGD1. According to the invention, three self-isolated, identified and preserved strains of H3N2 canine influenza virus CGD1, CGD2 and CGD3 are used to inoculate chick embryo for subcultring, and the CGD1 is found to be good in genetic stability. Therefore, the CGD1 is selected for plaque purification to breed an H3N2 canine influenza virus vaccine strain CIVGDYM1, and the vaccine strain has been preserved in China general microbiological culture collection center, with an accession number being CGMCCNO: 7218. By using the H3N2 canine influenza virus vaccine strain CIVGDYM1 to inoculate the chick embryo for virus breeding, and through gaining allantoic fluid of the chick embryo, inactivating, preparing vaccine and other steps, the safe and effective H3N2 canine influenza virus inactivated vaccine can be prepared.

The invention belongs to the technical field of biology, and particularly relates to a tetravalent influenza virus subunit vaccine and a preparation method thereof. Each dose of the tetravalent influenza virus subunit vaccine contains H1N1, H3N2 and two types of B; and the tetravalent influenza virus subunit vaccine is prepared by virus inoculation, virus multiplication culture, allantoic fluid harvesting, clarification, inactivation, ultrafiltration concentration, cracking and ultracentrifugal purification, mixing, filtration sterilization, split charging and packaging, wherein the inactivation process comprises the following steps: firstly, adding a carboxymethyl glucan solution into monovalent virus harvesting liquid, and then adding formaldehyde for inactivation. According to the method disclosed by the invention, the content of free formaldehyde in the prepared tetravalent influenza virus subunit vaccine is reduced, the storage life of the vaccine is prolonged, and the antibody level after vaccine immunization is increased.

The invention discloses a method for producing an avian influenza vaccine by using a WAVE bioreactor, which comprises the following steps: (1) cell culture: inoculating cells onto a micro supporter of a cell culture bag, adding a cell culture medium, and shaking the cell culture bag to perform cell culture; (2) virus solution multiplication: settling the micro supporter, washing the cultured cells, inoculating with an avian influenza virus seed, adding a virus multiplication culture medium, and further performing cell culture, thus obtaining a virus solution; and (3) inactivating the virus solution, and emulsifying to obtain the avian influenza vaccine. The method disclosed by the invention optimizes the technological parameters in the process of culturing MDCK cells by using a WAVE bioreactor, effectively improves the MDCK cell culture efficiency, and finally increases the avian influenza vaccine production efficiency remarkably. The immunoprotection efficacy experiment proves that the avian influenza vaccine prepared by the method disclosed by the invention has exact immunoprotection efficacy for poultry.

A reovirus vaccine and its method for preparation belong to the technical sphere of druggist sundries and are used to solve the indigenous fowl reovirus infection vaccine problem. The separated reovirus is of CGMCC No.1713 as its preserved number and has been preserved in China microbial bacterial preservation management committee common microorganism center on May 16th, 2006. The said vaccine is prepared by steps including virus isolation from organ of diseased chicken, serum check, virus breeding and generation and cryodesiccation. The invention separates the virus from the diseased chicken with ARV infection of indigenous fowls, applies them in immune chicken, and produces immune response within 1 week and produces strong immunity within 2 weeks. The immune time is 2-2.5 months, and the detoxicating protecting rate during immune period is 94.8%-98.8%, which is explicitly higher than import vaccine protecting rate and can make up for the domestic defect of vaccine vacancy in reovirus affection.

The invention discloses a full-suspension culture method for avian influenza virus. The full-suspension culture method comprises the following steps: step 1, taking EB66 cells for growth culture; step2, when the density of the EB66 cells grows to be suitable for virus inoculation, utilizing a fed-batch fluid-replacement virus inoculation technology to inoculate avian influenza virus into the EB66cells and performing virus multiplication culture; step 3, 24h after virus inoculation, sampling every 12h to determine HV titer of the virus and obtaining and storing the virus when the virus HA titer reaches to the maximum to obtain cultured avian influenza virus. According to the full-suspension culture method disclosed by the invention, the EB66 cells are utilized to perform avian influenza virus culture, so that the defect that a lot of miscellaneous protein is prone to being introduced into when a traditional chick embryo culture technology is utilized for production is overcome, and occurrence of chicken immunity side reaction is effectively reduced; meanwhile, titer and purity of the cultured avian influenza virus are improved; furthermore, production quality of the avian influenza vaccine is improved.

A collecting method in insect viruses breed includes: 1) feeding three-year old larva by 500,000 polyhedrons/ml concentration for 3 days, 2) primary collecting larva after experimenting 7-8 days under 24deg.C environment,3) extracting virus and processing viral imagocide after reserving 20-30 days under 15deg.C homeothermal condition. It achieves high insect virus output, simple viral collecting process, electrical save.

The invention relates to the technical field of biology, in particular to a method for producing a human diploid cell encephalitis B inactivated vaccine. The method sequentially comprises the following steps of: resuscitating and subculturing a human diploid cell strain; culturing the human diploid cell strain by using a multi-stage bioreactor; inoculating an encephalitis B virus strain, and performing virus multiplication culture; harvesting a virus culture solution; and inactivating the virus culture solution, performing ultrafiltration, purifying, adding a glycoprotein protective agent, and preparing the vaccine. Human diploid cells are used as a cell matrix, so that the vaccine is safe and is free from exogenous factor pollution; the human diploid cells are cultured by the bioreactor through stage-by-stage linear amplification, so that the large-scale production of the vaccine with low cost, high quality, safety and stability is easy to implement; and the titer of the virus harvested solution is subjected to sampling inspection every day, so that the high expression of the harvested virus culture is ensured, and production quality is ensured.

The invention discloses a method for preparing an avian influenza virus and an inactivated vaccine thereof with Vero passage cells. According to the method disclosed by the invention, the Vero passage cells are adopted to replace chicken embryo to culture influenza virus, so that the problems of chicken embryo remnant and exogenous virus inflection are solved and the immunogenicity of the cultured virus is more stable. On the other hand, for avoiding the phenomenon that the cracking of HA is influenced because protease is acted by an inhibitor in a maintaining liquid to inactivate, pancreatinis coated with chitosan and then is added in a cell maintaining liquid, so that the pancreatin is slowly released in the maintaining liquid and the defect that the cracking of hemagglutinin is influenced because the pancreatin is inactivated in the reproduction process of viruses is overcome. In addition, the probability that the cells are polluted since the pancreatin is added for multiple timesis avoided. The virus cultured by the method disclosed by the invention is high in titer and favorable in stability and is suitable for large-scale vaccine production.

The invention provides a new hepatitis A virus JS-4 strain for preparing hepatitis A inactivated vaccine and a method for culturing the same. The strain is separated from dejecta of a patient infected with acute hepatitis A and is transmitted to a Vero cell for adapted culture; through neutralization tests and other methods, the strain is proved to be hepatitis A virus; in the adaptation process, the subculture cycle of early generation is 28 days; when the strain is transmitted to the tenth generation, the cycle is shortened to 21 days; the strain is continuously cultured for ten generations; the culture temperature is between 35 and 36 DEG C; the cycle for virus propagation is shortened from 28 to 21 days; the antigen titer can reach 1:640-1:1,280; and the infectious titer is between 106.67 and 107.50 CCID50/ml. The marmoset virulence test proves that the strain has weak virulence, is reliable in safety and has good immunogenicity and protective effect when the strain is used for producing the hepatitis A inactivated vaccine and is an ideal strain for producing the hepatitis A inactivated vaccine.