25 results about "Mdck cell" patented technology

The invention relates to the technical field of biology, in particular to an MDCK cell strain and application of the MDCK cell strain in preparation of influenza vaccines. The invention provides an MDCK cell strain, the preservation number of which is CCTCC NO: C202504. The cell strain not only can maintain high sensitivity and high yield to influenza viruses and ensure the yield and quality of vaccines, but also can remarkably improve the biological safety of cell matrix influenza vaccines due to elimination of tumorigenicity risk, and has a good application prospect. And a key technical support is provided for promoting domestic research and development and marketing of influenza vaccines based on cell culture.

The invention discloses an MDCK cell low-serum culture medium construction method based on KAN modeling and Bayesian optimization and application thereof.The MDCK cell low-serum culture medium construction method comprises the steps that firstly, key influence factors are screened out from numerous nutrient components through multi-factor experimental design, and then a high-precision nonlinear prediction model between the key factors and cell performance is established through KAN; based on the model, a Bayesian optimization algorithm is adopted for rapid optimization, and the optimal concentration of each component is determined. The cell line culture medium has universality and can be migrated and applied to culture medium development of other cell lines. According to the MDCK cell low-serum culture medium constructed through the method, the adding amount of fetal calf serum (FBS) is only 2%-4% (v / v), the formula is systematically optimized, efficient growth of cells can be directly supported, and pre-domestication is not needed. The culture medium has obvious effects on promoting MDCK cell proliferation, improving virus (such as influenza virus) titer and reducing production cost, and is suitable for large-scale production of vaccines.

The invention discloses a non-tumorigenic MDCK cell strain, a screening method and application. The non-tumorigenic MDCK cell strain is MDCK-CA027 (the preservation number is CCTCC (China Center for Type Culture Collection) NO: C2023329). The tumor formation rate of the cell strain is 0% when the cell strain is observed for at least 16 weeks in a nude mouse in-vivo tumor formation test, and the cell strain has good sensitivity to various influenza viruses (such as H1N1, H3N2, BY and BV). The invention also provides a method for screening the cell strain. The method comprises the following steps: preparing a single-cell suspension, carrying out limited dilution to obtain monoclone, carrying out enlarged culture, carrying out virus sensitivity detection, carrying out in-vitro tumorigenicity related detection (clone formation test / soft agar test), carrying out nude mouse in-vivo tumorigenicity test and the like. The non-tumorigenic MDCK cell strain is high in safety, can efficiently support influenza virus replication, can be safely and effectively applied to influenza vaccine production and influenza virus amplification, overcomes the tumorigenic risk of traditional MDCK cells, and has important scientific significance and application prospects.

This invention provides a method for constructing and applying MDCK cells stably integrated with disialotyltransferase, relating to the field of biotechnology. The construction method includes: infecting MDCK cells with lentiviral particles, followed by culture and screening to obtain MDCK cells stably expressing porcine ST3GAL1 and porcine ST6GAL1 genes; the lentiviral particles carry porcine ST3GAL1 and porcine ST6GAL1 genes; the nucleic acid sequence of the porcine ST3GAL1 gene is shown in SEQ ID NO.1; the nucleic acid sequence of the porcine ST6GAL1 gene is shown in SEQ ID NO.2. This construction method is simple and convenient, and the constructed MDCK cells can stably express porcine ST3GAL1 and porcine ST6GAL1 genes, effectively improving the culture efficiency of swine influenza virus, and thus can be used in the preparation of swine influenza virus.

The invention belongs to the field of new application of medicines, and particularly relates to application of 4-hydroxyderrisin in preparation of anti-influenza virus medicines. It is found for the first time that 4-hydroxyderrisin can significantly inhibit replication and internalization of influenza A virus H1N1 and has significant anti-influenza virus activity, the median inhibitory concentrations of 4-hydroxyderrisin in MDCK cells and A549 cells are 0.60 M and 0.23 M respectively, and the 4-hydroxyderrisin is low in cytotoxicity and high in safety. Experiments show that 4-hydroxyderrisin plays a role in the early stage of a virus life replication cycle, and plays an antiviral role by inhibiting virus internalization without affecting virus adsorption and neuraminidase activity. The medicine can be prepared into a solid or solution dosage form, is used for preventing and treating influenza, and has a good application prospect.

The invention discloses an application of a Wevir drug in preparation of a drug for resisting canine distemper virus. It is found that 20-100 [mu] M of pirentamivir and ombitasvir have no obvious toxic effect on MDCK cells, and 20 [mu] M of pirentamivir and ombitasvir can significantly inhibit canine distemper virus proliferation, which indicates that pirentamivir and ombitasvir can become a new choice for preparation of anti-canine distemper virus drugs. The pirentamivir and ombitasvir pharmaceutical composition is a new use of an old medicine, the pharmacokinetic data of pirentamivir and ombitasvir which are sold on the market are detailed, the safety is reliable, the development of the new use can be quickly performed for second-stage clinical evaluation, the research and development period is shortened, the development cost is saved, and resources can be utilized to the maximum extent.

The invention provides an MDCK (Madin-Darby Canine Kidney) cell and application thereof. The MDCK cell screened by the invention is preserved in China General Microbiological Culture Collection Center (CGMCC) on November 06, 2025, and the preservation number is CGMCC NO: 46717. The screened non-tumorigenicity MDCK cells can be used as a cell matrix for influenza vaccine production. The suspension technology is adopted, and compared with traditional wall attaching, the suspension technology has the advantages of being easy to produce in a large scale, higher in production efficiency and yield, better in process control and consistency, lower in production cost and free of introduction of serum animal source components.

Disclosed is an adapted Madin-Darby canine kidney cell line capable of suspension culture in the absence of serum, and a chemically-defined medium for culture of the adapted MDCK cell line. Further disclosed are culture methods for growing the adapted MDCK cell line and methods for producing a vaccine from the adapted MDCK cell line grown in the chemically-defined medium.

The invention discloses application of verbascoside in preparation of a product for preventing and treating influenza virus infection, and relates to the technical field of biological medicines. Through molecular docking screening, cytotoxicity experiments and antiviral activity verification, the application value of verbascoside in prevention and treatment of influenza virus infection is defined, and the technical effect is remarkable. In the aspect of safety, the verbascoside has no obvious influence on the survival rate of MDCK cells at the concentration of 200 mu M, the concentration of the verbascoside is far higher than the clinical potential medication concentration, and the verbascoside has no obvious cytotoxicity and good biocompatibility. In the aspect of antiviral activity, verbascoside shows an efficient inhibition effect on influenza viruses (WSN strains and FM1 strains), the IC50 value is as low as 28.3 mu M, and the inhibition effect is superior to that of part of natural components. In an application scene, verbascoside can be prepared into various drug dosage forms and can also be used as an active ingredient of a disinfectant, and a novel safe, efficient and multifunctional flu prevention and control scheme is provided for the field of biological medicine.

The invention belongs to the technical field of biology, and discloses an influenza B virus sensitizing cell strain as well as a construction method and application thereof. The influenza B virus sensitizing cell strain STX-BB2 can be used for stably expressing influenza virus receptor ST6GAL1 protein in a human source and controllably inducing and expressing influenza B virus PB2 protein by using doxycycline; the preservation number of the cell strain STX-BB2 is GDMCC (China General Microbiological Culture Collection Center) NO: 67100; the cell strain can enhance the expression ability of a host receptor and the virus replication key gene level at the same time; the infection efficiency and the replication titer of the influenza B virus in MDCK cells can be improved; the method can be widely applied to influenza B virus separation, mechanism research, vaccine seed virus amplification and drug screening model construction.

The invention provides application of levistilide A in preparation of anti-influenza virus drugs, and belongs to the technical field of biological medicines. The influenza virus is an H1N1 virus. In-vitro anti-influenza experiments find that levistilide A can significantly inhibit infection of H1N1 virus and significantly improve cytopathy caused by influenza virus infection. The H1N1 virus half inhibitory concentration (IC50) of the levistilide A in MDCK cells is 1.462 M, and the H1N1 virus half inhibitory concentration (IC50) of the levistilide A in A549 cells is 0.455 M. The invention proves that the levistilactone A has a remarkable anti-H1NA virus effect, can be used for preparing the anti-influenza virus medicine, and has an excellent application prospect.

Provided are an FLNB gene-knockout MDCK cell line, a construction method therefor and the use thereof. The FLNB gene-knockout cell line is named MDCK-FLNB-KO, and has been deposited in China Center for Type Culture Collection (CCTCC) on November 20, 2024, and the accession number is CCTCC NO: C2024391. The FLNB gene knockout uses CRISPR / Cas9 technology, wherein a sgRNA is designed for exon 1 of the FLNB gene of MDCK cells, an FLNB gene knockout vector plasmid 1 and plasmid 2 are constructed, and the plasmid 1 and the plasmid 2 are transfected into the MDCK cells to obtain the FLNB gene-knockout MDCK cell line. Compared with the original MDCK cell line, the obtained cell line has a lower adhesion ability and a higher yield when applied to influenza virus culture.

This invention discloses a screening method, cell line, application, and cell bank for non-tumorigenic MDCK monoclonal strains, belonging to the field of biotechnology. The screening method includes: serum-free acclimation culture of a heterogeneous MDCK cell population; monoclonal isolation of the acclimated cell population to obtain candidate monoclonal cell lines; tumorigenicity screening by inoculating candidate monoclonal cell lines into immunodeficient animals to select non-tumorigenic cell lines; and viral susceptibility evaluation of the non-tumorigenic cell lines to select cell lines with predetermined susceptibility to a predetermined virus. This MDCK monoclonal cell line is non-tumorigenic. This invention addresses the risks of population heterogeneity and potential tumorigenicity in existing MDCK cell lines while ensuring viral production efficiency, providing an ideal cell platform for producing safer and quality-controlled viral vaccines.

This invention discloses a method for efficiently isolating human parainfluenza virus based on suspended MDCK cells. By using the fully suspended passaged MDCK cell line for human parainfluenza virus isolation and culture and optimizing the inoculation process, single-generation virus amplification can be completed within 3 days, significantly shortening the cycle, improving virus isolation efficiency and success rate compared to traditional adherent culture methods, and enhancing experimental reproducibility. This method not only demonstrates good isolation and culture capabilities for the major subtypes of human parainfluenza virus, but also shows good adaptability to clinical samples from different sources and with different initial loads. In addition, this culture system has linear scale-up potential, is not limited by the surface area of ​​adherent culture, and can meet the large-scale virus seed preparation needs from clinical testing to vaccine mass production, providing key technical support for the integrated process from human parainfluenza virus isolation to vaccine production.

The invention discloses application of long-chain non-coding RNA (LncRNA) XR002626937.1 in regulation and control of replication of canine coronavirus (CCoV), and belongs to the technical field of biological medicine. The nucleotide sequence of the LncRNA XR002626937.1 is as shown in SEQ ID No.1, is mainly positioned in a cell nucleus, is remarkably up-regulated in time and dose dependence after MDCK cells are infected by CCoV, and is positively correlated with virus load. Functional experiments prove that overexpression of the LncRNA can significantly promote CCoV replication, and knock-down of the expression of the LncRNA can effectively inhibit virus proliferation; the action mechanism comprises the steps of inhibiting activation of a p53 signal channel and downstream target genes p21 and Bax, repressing apoptosis of infected cells, antagonizing expression of natural immune factors such as RIG-I, MAVS, ISG15 and the like, promoting virus immune escape, and being capable of serving as competitive endogenous RNA to adsorb miR-10926-z to regulate and control the downstream signal channel. The key function of the LncRNA XR002626937.1 in CCoV infection is defined for the first time, a new target spot and a theoretical basis are provided for developing the anti-CCoV medicine targeting the LncRNA, and the application prospect is wide.

The invention discloses application of akebia quinata phenylethanoid glycoside B in preparation of a product for preventing and treating influenza virus infection, and relates to the technical field of biological medicines. Through molecular docking screening, cytotoxicity experiments and antiviral activity verification, the application value of the akebia quinata phenylethanoid glycoside B in prevention and treatment of influenza virus infection is defined, and the technical effect is remarkable. In the aspect of safety, the akebia quinata phenylethanoid glycoside B has no obvious influence on the survival rate of MDCK cells under high concentration, is far higher than clinical potential medication concentration, has no obvious cytotoxicity and is good in biocompatibility. In the aspect of antiviral activity, the akebia quinata phenylethanoid glycoside B shows an efficient inhibition effect on influenza viruses, the IC50 value is as low as 28.5 mu M, and the inhibition effect is superior to that of part of natural components and oseltamivir. In an application scene, the akebia quinata phenylethanoid glycoside B can be prepared into various drug dosage forms and can also be used as an active ingredient of a disinfectant, and a novel safe, efficient and multifunctional influenza prevention and control scheme is provided for the field of biological medicine.

The invention relates to the field of biology, and particularly provides a 293H cell line for expressing an influenza virus PB1 gene, the cell line is classified and named as an HEK-293H cell line, and the preservation number is GDMCC NO: 66761. The preservation date is July 25, 2025, and the strain is preserved in the Guangdong Microbial Culture Collection Center, and the preservation address is the 5th floor of the Experimental Building, No. 100 Courtyard, Middle Xianlie Road, Guangzhou District, Guangdong Province. In addition, the invention also provides an MDCK cell line for expressing the influenza virus PB1 gene, both the MDCK cell line and the MDCK cell line can express PB1 protein, and rescue and propagation of the PB1 gene knockout (PB1-KO) replication defective influenza virus can be realized. Meanwhile, the invention also discloses application of the two cell lines.

The application discloses a QMC influenza virus seed, a preparation method and application thereof, and belongs to the technical field of biological medicines.The preparation method of the QMC virus seed is virus passage adaptation and virus seed preparation which are synchronously performed, and comprises the following steps: inoculating chicken embryo source influenza virus on MDCK-XF06 cells, and obtaining recovered passage virus seed after culture; then inoculating the recovered passage virus seed on MDCK cells according to a mode of adjusting virus inoculation infection multiplicity from generation to generation, and obtaining P2 generation master seed batch virus seed and P3 generation working seed batch virus seed after culture.The QMC virus seed obtained through the preparation method has the following advantages: good virus activity, high virus titer, efficient production of a large amount of live virus, excellent genetic stability, ensured consistency and effectiveness of product quality, completely eliminated tumorigenicity risk, high safety, and the like, and lays a solid foundation for subsequent production of safe, efficient and stable vaccine products.

The invention discloses a method for full-suspension serum-free efficient production of canine distemper virus by adopting an MDCK cell line and application, and belongs to the technical field of virus amplification. In order to overcome the technical defects existing in production of the canine distemper virus by using adherent growth chick embryo fibroblasts or Vero cells (E6), the invention provides a method for producing the canine distemper virus by inoculating the canine distemper virus into an MDCK (Madin Darby Canine Kidney) cell serum-free full-suspension cell line and using a bioreactor, and the method is subjected to process optimization, so that the canine distemper virus can be produced. According to the optimized method, the production cost is remarkably reduced, the production period is short, the virus content is high, the oxygen transfer coefficient in the culture process is increased, the production time consumption is reduced, and the virus production efficiency and the vaccine yield and quality are improved. The method provided by the invention can meet the large-scale efficient production of the canine distemper virus, and has a better application prospect in the production of canine distemper virus vaccines.

The present disclosure provides compositions and methods for manufacturing and obtaining a live attenuated Influenza vaccine (LAIV) composition that can be delivered intranasally to provide protection against influenza virus infection. Said LAIV strains are based on cold adapted, temperature sensitive and attenuated phenotypes of master donor viruses (MDVs) containing the surface glycoprotein genes of the wild type pandemic or seasonal influenza strains. Also, said LAIV strains are further adapted to grow in MDCK cells (Madin Darby canine kidney cells). The use of eggs is avoided in large scale vaccine manufacturing. The purification process is devoid of chromatography steps. The said LAIV composition includes one or more live attenuated influenza vaccine virus and is devoid of polymers and surfactants.

This invention belongs to the field of nanobody technology and provides a method for preparing and applying a single-domain antibody against avian influenza virus H9N2. The application involves using this H9N2 single-domain antibody in the prevention and control of avian influenza virus H9N2. The invention prepares the H9N2 single-domain antibody by immunizing striped bamboo sharks and collecting their tail vein blood. The purity and stability of the H9N2 single-domain antibody were evaluated. The results show that the recombinant single-domain antibody targeting the H9N2 virus HA protein prepared in this invention has high purity and good activity, exhibits good in vitro binding activity with H9N2, and shows good stability at room temperature. The recombinant single-domain antibody 28-H9N2-2SP4H can recognize and bind to the H9N2 virus on the surface of MDCK cell membranes, providing a new direction for the prevention and control of avian influenza as an alternative to traditional antibodies.

The invention belongs to the technical field of biology, and discloses a preparation method of an MDCK stably transfected cell line, MDCK cells are co-transfected through a transposase expression plasmid and a transposase expression plasmid, and the MDCK stably transfected cell line integrated with a CAV2-E1 gene and capable of expressing an E1A gene and an E1B gene is obtained. The transposon plasmid comprises a CAV2-E1 gene and transposon elements which are added at the two ends of the CAV2-E1 gene. The cell line has relatively high rescue efficiency and multiplication capacity aiming at CAV-2, especially replication-deficient CAV viruses. Meanwhile, the invention further provides a preparation method and application of the MDCK stably transfected cell line, and further provides a method for rescuing replication-deficient CAV viruses by adopting the cell line.

The invention relates to the technical field of biomedical engineering, in particular to an MDCK cell strain with high yield of influenza virus and application of the MDCK cell strain, and particularly relates to construction of a cell cycle 20 protein (CDC20) knock-down stable cell line and application of the cell line in influenza virus proliferation. The invention provides an MDCK (Madin Darby Canine Kidney Cells) cell strain with high yield of influenza virus, the MDCK cell strain is a CDC20 knock-down stably transfected MDCK cell strain, the MDCK cell strain is named as MDCK-sh-cdc20 (Madin Darby Canine Kidney Cells), the MDCK-sh-cdc20 is preserved in China Center for Type Culture Collection (CCTCC), the preservation address is Wuhan University, Wuhan, China, the preservation number is C2025105, and the preservation date is 2025.4.22. The invention further provides a preparation method of the MDCK cell strain. The invention further provides a construction method of the MDCK cell strain, and the CDC20 knock-down stably transfected MDCK cell strain is obtained by mediating silencing of CDC20 genes in MDCK cells through an RNAi technology. The invention also provides an application of the MDCK cell strain in multiplication culture of influenza virus.

This invention relates to the field of bioengineering, and more particularly to sgRNA, MDCK cells, and their applications. This invention provides sgRNA having the nucleotide sequences shown in SEQ ID NO:1 to SEQ ID NO:4. This invention employs a CRISPR-Cas9 gene editing method, which, compared to existing RNAi methods, offers higher specificity and lower off-target risk. Compared to MDCK cells, PCR detection showed a reduction of over 90% in RNA levels in IRF7- / - cells, Western blotting showed a significant decrease in IRF7 protein expression, and viral hemagglutination assays showed higher titers for influenza virus compared to the original cells. Tumorigenicity assays showed that the gene knockout cells retained low tumorigenicity. Compared to existing technologies, the cells of this invention exhibit both low tumorigenicity and significantly increased viral yield, making them suitable for influenza vaccine production.

The invention provides a composition useful to prepare high titer influenza B viruses, e.g., in the absence of helper virus, which includes internal genes from an influenza B virus vaccine strain or isolate, e.g., one that is safe in humans, for instance, one that does not result in significant disease, that confer enhanced growth in cells in culture, such as MDCK cells, or in eggs.