665 results about "Cell-cell fusion" patented technology

The invention relates to a method for separating mesenchymal stem cells from placenta. The method comprises the following steps: (a) taking placental cotyledon, and fully washing by using a phosphate buffer solution (PBS) to remove residual blood from the placenta; (b) cutting the placental cotyledon into blocks, adding a PBS containing tissue digestive enzyme, and incubating and digesting at 37DEG C; (c) filtering the tissue blocks by using a copper net, and grinding if necessary to promote filtration; (d) centrifuging the collected filtrate, separating mononuclear cells, suspending the obtained cells by using a mesenchymal stem cell (MSC) culture medium, and culturing in a 5 percent CO2 incubator at 37DEG C; and (e) after the dispersed cells form clones, selecting the clone cells, respectively culturing by using an MSC culture medium, and after the cells are fused, performing digestion and passage by using pancreatin to obtain the mesenchymal stem cells of the placenta. By the method, high purity mesenchymal stem cells of the placenta can be obtained.

The present invention relates to a method of preventing or inhibiting viral infection of a cell and/or fusion between the envelope of a virus and the membranes of a cell targeted by the virus (thereby preventing delivery of the viral genome into the cell cytoplasm, a step required for. viral infection). The present invention particularly relates to the families of RNA viruses, including the arenaviruses, coronaviruses, filoviruses, orthomyxoviruses, paramyxoviruses, and retroviruses, having Class I membrane fusion proteins as the fusion proteins that mediate this fusion process. The present invention provides for a method of identifying a conserved motif or domain called the fusion initiation region (FIR) in these viruses. The present invention further provides for methods of preventing infection by such viruses, by interfering with their FIR. The present invention further provides for methods of treatment and prophylaxis of diseases induced by such viruses.

A group of compounds that inhibit HIV replication by blocking HIV entry was identified. One representative compound, designated NB-206, and its analogs inhibited HIV replication (p24 production) with IC₅₀ values at nanomolar levels. It was proved that NB-206 and its analogs are HIV entry inhibitors by targeting the HIV gp41 since: 1) they inhibited HIV-mediated cell fusion; 2) they inhibited HIV replication only when they were added to the cells less than one hour after virus addition; 3) they blocked the formation of the gp41 core that is detected by sandwich enzyme linked immunosorbent assay (ELISA) using a conformation-specific MAb NC-1; and 4) they inhibited the formation of the gp41 six-helix bundle revealed by fluorescence native-polyacrylamide gel electrophoresis (FN-PAGE). These results suggested that NB-206 and its analogs may interact with the hydrophobic cavity and block the formation of the fusion-active gp41 coiled coil domain, resulting in inhibition of HIV-1 mediated membrane fusion and virus entry.

The invention relates to proteins comprising HIV fusion inhibiting peptides, such as T-20 and/or T-1249 peptides (including, but not limited to, fragments and variants thereof), which exhibit anti-retroviral activity, fused to albumin (including, but nbot limited to fragments or variants of albumin). These fusion proteins are herein collectively referred to as “albumin fusion proteins of the invention.” These fusion proteins exhibit extended shelf-life and/or extended or therapeutic activity. The invention encompasses therapeutic albumin fusion proteins, compositions, pharmaceutical compositions, formulations and kits. The invention also encompasses nucleic acid molecules encoding the albumin fusion proteins of the invention, as well as vectors containing these nucleic acids, host cells transformed with these nucleic acids and vectors, and methods of making the albumin fusion proteins of the invention using these nucleic acids, vectors, and/or host cells. The invention also relates to compositions and methods for inhibiting HIV-induced cell fusion. The invention further relates to compositions and methods for inhibiting HIV transmission to uninfected cells.

The invention relates to a separation method and a culture method for umbilical cord mesenchymal stem cells. The separation method comprises the following steps: thoroughly cleaning umbilical cord tissue of a healthy newborn by using a PBS (phosphate buffer solution) containing streptomycin and penicillin, and removing blood; shearing the umbilical cord into small sections uniform in length, and mechanically separating, bluntly stripping Wharton' s jelly, and removing umbilical arteries and umbilical veins; uniformly shearing the Wharton' s jelly; re-suspending the sheared Wharton' s jelly through an MSCs (mesenchymal stem cells) culture medium, inoculating to a culture dish with laid gelatin, and putting in a CO2 culture box for cultivation; conducting centrifugal separation to obtain tissue blocks and a cell resuspension solution. The culture method comprises the following steps: enwrapping the culture dish, discarding the gelatin, and washing with the PBS; inoculating the separated out tissue blocks and the cell resuspension into the culture dish; performing digestive subculture after cell fusion growth rate reaches 80-90%.

The invention relates to a assaying method for the aflatoxin, specifically saying, it is a method of adopting immunity affinity column and fluorophotometer prepared by monoclonal antibody of aflatoxin to assayt the aflatoxin in the food, which belongs to biocytology and microbacteria toxic metabololic products assaying method technical field.The method is as following: first, preparing monoclonal antibody by the steps involving: immunizing mouse, cell fusion, screening crossing tumour and obtaining the cell line of the crossing tumourc, gathering monoclonal antibody, etc, then using the abovementioned monoclonal antibody to prepare the immunoaffinity column, last, after milling the food and feed, extracting the aflatoxin out of it with methanol/water solution, after filtering, letting the liquid sample flow through the affinity column to be refined, successively, eluting the AFT from the affinity column with methanol, and assaying the content of aflatoxin in the sample with a fluorophotometer.

The invention provides methods for fusing a first cell with a second cell to form a hybrid cell. The methods involve incubating a first parental cell producing a first partner of a fusogenic binding partner pair on its surface with a second parental cell producing a second partner of the fusogenic binding partner pair on its surface. In certain embodiments, the parental cells are incubated with a known fusogen such as polyethylene glycol and the fusogenic binding partner pair increases the rate of cell fusion. In many embodiments, the first cell is an antibody producing cell, the second cell is an immortal cell, and the hybrid cell is a hybridoma cell that produces a monoclonal antibody. Also provided by the invention are methods for producing hybridoma cells, and methods for screening those cells for production of a monoclonal antibody of interest. The invention further provides systems and kits for carrying out the subject methods. The subject methods, systems, and kits find use in a variety of different industrial, medical and research applications.

The invention discloses a hybridoma cell strain capable of secreting monoclonal antibodies to quinolones and application of the monoclonal antibodies thereof. Ciprofloxacin (CIP) coupled with bovine serum albumin is used as an antigen to immunize BALB/c mice and cell fusion, screening and cloning are carried out so as to obtain one hybridoma cell strain 1F1 capable of stable passage and secretion of monoclonal antibodies (MAb) to quinolones, wherein, the accession number of the hybridoma cell strain 1F1 is CGMCC No. 5608. The titres of ascitic fluids of the 1F1 monoclonal antibodies are up to 10<-7>, and the type and the subclass of the monoclonal antibodies are IgG1 and kappa chain. According to indirect competitive ELISA analysis, the 1F1 monoclonal antibodies perform specific reactions to quinolones like ciprofloxacin, enrofloxacin, ofloxacin, danofloxacin, norfloxacin, enoxacin, marbofloxacin, sarafloxacin and difloxacin. An ELISA method, a kit and test paper for detecting residual of quinolones in food are developed by using the 1F1 monoclonal antibodies.

The invention relates to a method of realizing the screening and the monoclone preparation of hybridoma cell in one step. The method comprises that the semisolid medium of the monoclone of hybridoma cell is firstly prepared, the basic fibroblast growth factor of human is added into the fusion cell, then hydridoma fusion factor and clone factor are added and well-mixed, then the mixed material and the semisolid medium are gently well-mixed and packaged into plates and cultivated in the condition of 35 DEG C and CO2 with concentration of 5 percent; when visible clone colony grows in the plates, the clone is aspirated from the culture medium by a micropipettor and moved onto a cell culture plate for liquid scale-up culture, and one clone is moved in one hole; till the cell grows to half to two-third of the hole bottom, the culture supernatant is aspirated to do positive detection, and the positive hole is chosen to directly scale-up frozen and stored for further carrying out the characteristic identification of antibody or ascites production. The method of realizing the screening and the monoclone preparation of hybridoma cell in one step has the advantages of the simple operation, the short culture period, not requiring feeder cells and easily achieving the screening and the monoclone preparation of hybridoma cell after cell fusion in one step.

The invention provides a method for separating and culturing umbilical cord blood mesenchymal stem cells. The method comprises the steps of carrying out secondary separation on umbilical cord blood mononuclear cells by using a separating medium, and then inoculating the umbilical cord blood mononuclear cells into a culture bottle in which fibronectin and CD90 monoclonal antibodies are coated; and culturing for 4-5 days by using a serum-free culture system and simulating the in-vivo low-oxygen growth environment of the mesenchymal stem cells, then, removing suspension cells, further culturing adherent cells, and subculturing after the fusion rate of primary cells is up to 60%. By using the method for separating and culturing umbilical cord blood mesenchymal stem cells, provided by the invention, the problems of adherence infirmness, low culture success rate, low purity and the like caused in the extraction and culture processes of the umbilical cord blood mesenchymal stem cells are effectively solved, and the safety in clinical application is improved. In addition, the application ranges of the umbilical cord blood mesenchymal stem cells are widened, the utilization values of the umbilical cord blood mesenchymal stem cells are developed, and the clinical application prospects of the umbilical cord blood mesenchymal stem cells are widened.

The invention discloses a sorghum mosaic virus tri-anti sandwich enzyme-linked immuno sorbent assay (TAS-ELISA) kit which comprises positive control, negative control, a sorghum mosaic virus polyclonal antibody, a sorghum mosaic virus monoclonal antibody, an enzyme labeled antibody and other materials and drugs, wherein the sorghum mosaic virus polyclonal antibody is obtained through rabbit immune separation serum after separating and purifying a sorghum mosaic virus sugarcane separator; and the sorghum mosaic virus monoclonal antibody is obtained by mouse immune, cell fusion and clone purification. The TAS-ELISA kit prepared by the invention has high detection sensitivity, strong specificity, high repeatability and high economic usefulness.

The invention discloses a monoclonal antibody against virulent strain E2 protein of classical swine fever virus and a hybridoma cell strain secreting the monoclonal antibody. The hybridoma cell strain is obtained by using hog cholera lapinized virus vaccine strain E2 protein expressed by Baculovirus as tolerogen, selecting Shimen strain E2 protein as immunogen, immunizing mouse by cyclophosphamide immunosuppression method, carrying out cell fusion, and sieving hybridoma cell strain capable of stably secreting monoclonal antibody against E2 protein. The monoclonal antibody can react with Shimen strain and can produce specific reaction with virulent strain of classical swine fever viruses of 1.1, 2.1, 2.2 and 2.3 gene sub-groups. The monoclonal antibody has neutralization activity and does not react with hog cholera lapinized virus vaccine strain, so that the monoclonal antibody can be used for differentiating virulent strain of classical swine fever virus and hog cholera lapinized virus vaccine strain, which establishes the foundation for establishing a method for differentiating wild virus infection of classical swine fever and vaccine immunity and for researching the molecular difference between CSFV virulent strain and mild strain.

The invention relates to a kit used for rapidly detecting Escherichia coli O157:H7 in a sample, and a detection method thereof. The invention belongs to the technical field of immunological detection. According to the invention, a heated and deactivated Escherichia coli O157:H7 immunogen is used for immunizing a healthy New Zealand rabbit, such that a polyclonal antibody is obtained, and the polyclonal antibody is adopted as a coating antibody; a BALB/C mouse is immunized, and cell fusion is carried out, such that a monoclonal antibody is obtained, and the monoclonal antibody is adopted as a secondary antibody; and a double-antibody sandwich ELISA kit of Escherichia coli O157:H7 in foodstuffs (meat) is established. With the kit, a rapid and highly efficient detection means is provided for the detection of the residue of Escherichia coli O157:H7 in foodstuffs. The kit is advantaged in relatively low cost, relatively good stability, and relatively good repeatability. According to the invention, a detection limit is 105cfu/mL. The kit and the method are suitable for large-batch detections of samples.

The invention relates to an anti-rabbit hemorrhagic disease virus (RHDV)VP60 albumen monoclonal antibody, and belongs to the technical field of biology. An SP2/0 myeloma cell and a BALB/c mouse splenic cell immunized by utilizing RHDV to recombine VP60 albumen undergo cell fusion, are selectively cultured by an HAT culture medium and undergo double ELISA screening by utilizing the recombined VP60 albumen and RHDV; the obtained cell culture supernatant is checked up and screened respectively to obtain a hybrid tumor cell strain A3C which can stably excrete the anti-RHDV VP60 albumen monoclonal antibody; the ascitic fluid ELISA titer of the A3C is detected to be 1:327,600; and according to identification, the monoclonal antibody can specifically combine the expressed recombined VP60 albumen as well as the RHDV, and one single reaction strip appears in both specific combinations, thereby proving that the anti-RHDV VP60 albumen monoclonal antibody is a VP60 specific antibody of the RHDV capsid albumen.

The invention belongs to the technical field of rape molecular breeding, and specifically relates to a breeding method of cabbage type rape woad oil cytoplasm male sterile line. The invention also relates to a protoplast fusion technology to recombinate the cytoplasm and nuclear genome of cabbage type rape-woad so as to establish cytoplasmic male sterility of recombinant cytoplasm. The breeding process comprises the following steps: obtaining the leaf meat protoplast of cabbage type rape-woad through separation and extraction, obtaining the protoplast fused somatic cell hybrid of cabbage type rape-woad through a cell fusion method; taking the obtained somatic cell hybrid of cabbage type rape-woad and leaf meat protoplast cell hybrid of cabbage type rape-woad as the female parent, taking cabbage type rape as the male parent, carrying out multi-generation backcross, and obtaining cytoplasmic male sterile line with carpelloid stamen; wherein the mitochondrion gene PCR amplification verifies that mitochondrial genome carries out recombination during the fusion process, thus the nucleoplasm becomes uneven, and the sterile character appears.

The invention discloses a method for cultivating triploid dendrobium huoshanense by a crossbreeding method of tetraploid plants and diploid plants. The method comprises the following steps: (1) dendrobium huoshanense species which are from different producing areas and have characteristics of good stress resistance, rapid growth, thick stalk, long stem, many tillers, low fiber content and high active ingredient content are preferably selected as polyploidy cultivated original plant parents; (2) seeds of the different species of original plants undergo aseptic sowing in a solid medium so as to obtain diploid protocorms; (3) protoplasts of the different species of diploid protocorms are prepared and cell fusion is carried out; (4) detected tetraploid plants undergo asexual tissue culture so as to obtain tetraploid plant seedlings; (5) the tetraploid plant seedlings undergo hardening-seedling acclimatization and cultivation to flowering; (6) the tetraploid plants and diploid plants undergo artificial pollination and crossbreeding, and fructification is completed to obtain triploid dendrobium huoshanense seeds; and (7) the triploid seeds undergo aseptic sowing and culture so as to obtain a lot of triploid seedlings.

Provided herein are a hybridoma cell line producing monoclonal antibody against foot-and-mouth disease virus (FMDV), the monoclonal antibody therefrom, reagent and kit for ELISA, and immunoassay method. The hybridoma cell line is produced by cell fusion of a parental cell and a myeloma cell line and has the same characteristics as the cell line whose strain designation is CmA40 and deposition number is ATCC (To be Provided). The parental cell is a splenocyte isolated from the spleen of a mouse immunized by an antigen derived from a 3ABC non-structural protein (NSP) of FMDV. The antigen used here is expressed by a prokaryotic cell. The monoclonal antibody produced by the hybridoma cell line can specifically recognize a 3ABC polypeptide and does not cross-react with an antiserum of swine vesicular disease virus.

The invention discloses a preparation method of deciduous tooth mesenchymal stem cells, comprising the steps of: cleaning and sterilizing the tissue surfaces of deciduous teeth with normal saline and 75% alcohol, preserving the deciduous teeth in deciduous teeth preserving fluid; acquiring dental pulp from the deciduous teeth, adding dental pulp digestive fluid to the dental pulp for digestion; obtaining a unicell suspension; adding cell culture fluid, carrying out centrifuging and cleaning, adding precipitated cells into cell culture fluid, putting the cell culture fluid in a carbon dioxide incubator for culturing; when the primary cell fusion reaches 70%, adding cell washing fluid, shaking the culture bottle for washing cells, sucking and abandoning the cell washing fluid, adding cell digestive fluid for digestion, adding cell culture fluid to terminate digestion, repeatedly beating upon the bottle bottom until the cells completely fall off, adding cell culture fluid, inoculating into a culture bottle, putting the culture bottle in the carbon dioxide incubator for culturing, regarding the cells as P1 generation mesenchymal stem cells; when the P1 generation cell fusion reaches 70-80%, carrying out trypsin digestion, collecting digestive cells, carrying out centrifuging and inoculation; after 3d, carrying out trypsin digestion again, collecting digestive cells, carrying out centrifuging, counting and inoculation, and when the P3 generation cell growth reaches 80%, gathering and cryopreserving the cells.