614 results about "Serum-Free Culture Media" patented technology

This invention relates to a method for amplifying neural stem cells in vitro by 3-dimensional culture. The method comprises: selecting microcarrier with 3-dimensional environment, pre-treating, coating the microcarrier with 40-60 ng/mL alkaline fibroblast growth factor, 40-60 ng/mL epidermal growth factor, and B27 DMEM/F12 neural stem cell serum-free culture medium, adding 1X105-1X106 neural stem cells into the culture bottle, taking out the microcarrier grown with neural stem cells, removing the microcarrier, and rinsing cells to obtain neural stem cells. The porous microcarrier can enlarge the culture area. The alkaline fibroblast growth factor and epidermal growth factor can promote cell multiple fission and improve cell microenvironment, which is advantageous for multiple fission of neural stem cells.

The invention relates to the field of biology, and discloses a serum-free culture medium which essentially comprises an IMDM (Iscove Modified Dulbecco Medium), L-glutamine, sodium bicarbonate, Hepes, recombinant human insulin, recombinant human transferrin, recombinant human albumin, 2-mercaptoethanol, protocatechuic acid, lipid, amino acid, vitamins, trace elements, Pluronic F-68, hydrocortisone, vitamin C, bonding amine or recombinant human fibronectin, progesterone, putrescine, heparin, serotonin, epidermal growth factors (EGFs), b-fibroblast growth factors (FGF), platelet derive growth factor (PDGF)-BB and insulin-like growth factor (IGF)-I. The serum-free culture medium is clear in chemical components, free from animal sources and serum and safe and ideal in cell cultivation and avoids the doped animal components and unstable batches, and the results of the cultured mesenchymal stem cells show that the total cellular score, the cell phenotype and the secretory cell factors are normal, so that the serum-free culture medium has good industrial application prospect.

The invention relates to a human amnion mesenchymal stem cell serum-free culture medium and a culture method thereof. The culture medium is formed by adding human serum albumin, human transferrin, human insulin and sodium selenite into a DMEM/F12 basic culture medium. The culture method for the culture medium comprises the following steps of: digesting human amnion by using trypsin, then digesting the human amnion by using collagenase IV and deoxyribonuclease I, and filtering the mixture to obtain single cell suspension; and adding the human serum albumin, the transferrin, the insulin and the sodium selenite into the DMEM/F12 basic culture medium in a ratio of VDMEM to VF12 of 1:1, and putting human amnion mesenchymal stem cells in a 37 DEG C CO2 incubator with saturated humidity and volume fraction of 5 percent under the serum-free condition, wherein culture in vitro and amplification are realized by solution change and transfer of culture, potentiality of multi-direction differentiation is maintained, and the amplified cells can be induced in vitro to form cartilage cells, osteoblasts and adipocytes. The culture medium and the culture method have the characteristics of no other animal sources, wide source and no limitation of ethics.

The invention provides a technological flow with versatility and high efficiency for reforming human serum albumins and fusion proteins of human serum albumins by separating and purifying target proteins. The steps of the method are as follows: the inorganic salt culture medium is used to acquire the genetic engineering microzyme fermented supernatant containing target proteins, and the serum-free culture medium is used to acquire a culture solution of reforming vertebrate cells, and ion exchange type affinity column chromatography medium Capto MMC fillers with higher salinity resistance and high capacity are directly used for separation and purification. The technological flow of the invention has the advantages of simple purification program step, uniqueness, low costs and convenient industrialization.

The invention discloses a serum-free medium for Madin-Darby canine kidney (MDCK) cell full suspension culture. The serum-free medium for MDCK cell full suspension culture comprises an amino acid part, a vitamin part, an inorganic salt part and other additive parts. The serum-free medium has the beneficial effects that the serum-free medium for MDCK cell full suspension culture, provided by the invention, is high in cell culture density and clear in composition and does not contain animal serum, a downstream product is purified, the product quality is improved, and the serum-free medium is convenient to prepare and use and suitable for large-scale production of influenza vaccines and avian influenza vaccines.

The invention relates to the field of cell engineering and particularly relates to a method for separating human adipose-derived stem cells from human adipose tissues. According to the method, by adding a small amount of low-molecular-weight heparin calcium into normal saline, a mixed solution is obtained and used as a cleaning fluid to clean the human adipose tissues, residual impurities such as red blood cells in the human adipose tissues can be well removed and thus human adipose-derived stem cells with relatively higher purity can be separated, the content of the parenchyma cell is significantly reduced, the differentiation induction of the human adipose-derived stem cells is facilitated, the consumption amounts of collagenase I, trypsase and a serum-free culture medium can be reduced and the cost of the process is also decreased. In addition, by centrifuging and digesting the human adipose tissues for 1 hour at a speed of 150r/min, the human adipose-derived stem cells with significantly better activity can be obtained.

The present invention provides a method for expanding an endothelial progenitor cell in vitro. More particularly, the present invention provides a method for culturing a hemangioblast comprising incubating a hemangioblast in a serum-free culture medium containing one or more factors selected from the group consisting of stem cell growth factor, interleukin-6, FMS-like tyrosine kinase 3 and thrombopoietin, and a vascular endothelial cell produced by the method; and a serum-free culture medium containing one or more factors selected from the group consisting of stem cell growth factor, interleukin-6, FMS-like tyrosine kinase 3 and thrombopoietin, and a kit for the preparation of the serum-free culture medium and the like.

The invention discloses a human-derived adipose tissue-derived stem cell factor, a preparation method and application thereof, and particularly discloses a method for culturing a human-derived adipose tissue-derived stem cell factor. The method comprises the following steps: culturing human adipose tissue-derived stem cells in a high-coenzyme serum-free culture medium until the cell confluence is 75-85 percent, and collecting supernatant. The invention further discloses a method for preparing human-derived adipose tissue-derived stem cell factor concentrated solution, the concentrated solution prepared by the method, further application of the concentrated solution, a cell factor concentrate prepared by the method, cosmetics and medicines prepared from the cell factor concentrate. The concentrated solution prepared by the method has high content of adipose tissue-derived stem cell factors and stable quality, and a product prepared by combining the concentrated solution with other materials in a specific ratio has a good effect of repairing skin injury; and the method is simple and feasible, is suitable for large-scale production, and has excellent market prospect.

The invention relates to a method for culturing gamma-delta-T cells, and in particular relates to a method for in-vitro amplification of gamma-delta-T cells, wherein the method comprises the following operating steps of: pre-coating a T75 culture bottle by a TCR-gamma-delta resisting antibody and CD28McAb for later use use; isolating the peripheral blood mononuclear cell (PBMC) of a patient; regulating the PBMC concentration to 1*10<6> 6/ml by a serum-free culture medium which contains 5% of autologous plasma, and transferring PBMC cell suspension into the T75 culture bottle; adding an initial culture medium which contains proper concentrations of Zoledronat, HSP70, 1L-2, 1L-7 and 1L-15; culturing in a saturated humid environment containing 5% of CO2 at 37 DEG C; depending on growth situation of the cell, changing the culture medium every 2-3days, to control the cell concentration at about 2.5*10<6>/ml; meanwhile, compensating full doses of Zoledronat, HSP70, 1L-2, 1L-7 and 1L-15; and continuously culturing for 12-16days, to obtain a great amount of gamma-delta-T cells which are comparatively high in purity.

The invention relates to a method for amplifying cytokine induced kill (CIK) cells and a CIK cell preparation, which belong to the field of in-vitro culture of immune cells. The method concretely adopts the following procedures that: a, lymphocyte cell separation liquid is used for separating out peripheral blood mononuclear cells (PBMC), a culture bag is covered by CD3mAb and CD137mAb in advance, the concentration of the PBMC obtained through separation is regulated to 1*10<6>/ml by a serum-free culture medium, in addition, IFN-gamma is added to obtain the final concentration being 1000 mu/ml, and the materials are transferred to the culture bag to be cultured; b, CD3mAb, CD28mAb and CD137mAb are added after the culture for 24h, in addition, the prepared serum-free culture medium is added, IL-1alpha, IL-2, IL-12 and IL-15 are added into the prepared serum-free culture medium, and obtained CIK cells are collected through centrifugation after the continuous culture for 7 to 21 days; and c, in the culture process of the step b, the cells in the culture bag are counted every three days, in addition, the culture medium is supplemented according to the concentration of the cells, and the CD3mAb, the CD28mAb and the CD137mAb are added to the corresponding concentration every six days, so the CIK cell generative cell times and the cytotoxin activeness are improved.

The invention provides BHK-21 cells obtained by high-density suspension culture in a low-serum and serum-free culture medium and a culture method thereof. The culture method comprises the following steps: 1) resuscitating frozen BHK-21 cells and performing adherent culture to obtain adherently cultured BHK-21 cells; 2) subculturing the adherently cultured BHK-21 cells in culture solution to obtain suspension-cultured BHK-21 cells and freezing the BHK-21 cells; 3) performing resuscitation culture of the suspension-cultured BHK-21 cells and performing biological property identification; and 4) performing bioreactor adaptive culture of the suspension-cultured BHK-21 cells identified to be qualified. The invention has the advantages that: a cell domesticating method provided by the invention and the obtained BHK-21 cells which can be cultured in a suspended manner save culture solution and bovine serum, are in accordance of current energy-saving, environment-protection and low-carbon concepts for the application of suspended-cultured cells into actual production saves floor area and labor investment greatly, and create considerable economic benefit and social benefit.

The invention relates to the cell culture technical field, and particularly relates to a natural killer cell culture medium and a natural killer cell amplification culture method. The invention provides the culture medium used for amplification culture of natural killer cells and containing a serum-free culture medium, human plasma, IL-2, IL-21, IL-15 and OKT-3. The culture medium provided by the invention is used for amplification culture of the natural killer cells, can avoid risks caused by exogenous serum, has high amplification efficiency and allows the obtained natural killer cells to have high purity. With adopting the method for amplification of the natural killer cells, amplification of the natural killer cells can be maintained at a logarithmic phase for a longer period of time. Moreover, the cultured natural killer cells have good killing activity on tumor cells.

The invention provides a method for producing a recombinant polypeptide of interest which method comprises: (a) providing a host cell which comprises a nucleotide sequence which encodes the recombinant polypeptide of interest and which directs expression of the recombinant polypeptide of interest in the host cell; (b) providing a serum-free culture medium which comprises (i) water, a plant-derived peptone, an osmolality regulator, a buffer, an energy source, at least one amino acid, a lipid source or precursor, a source of iron, non-ferrous metal ions and optionally one or more vitamins and cofactors; and (ii) does not contain any full-length polypeptides; and (c) culturing the host cell in the culture medium under conditions that allow for expression of the recombinant polypeptide of interest.

Pluripotent cells are derived and maintained in a self-renewing state in serum-free culture medium comprising a MEK inhibitor, a GSK3 inhibitor and an antagonist of an FGF receptor.

The invention discloses a serum-free culture medium without animal origin components for culturing a Vero cell micro-carrier. The serum-free culture medium consists of a DMEM/F12 (1:1) culture medium and culture medium additive components such as epidermal growth factors, insulin, serotonin, aurin tricar-boxylic acid, biotin, vitamin C, amino acid, fructose, trehalose, trace elements, hydroxypropyl-beta-cyclodextrin and the like. The serum-free culture medium has the following advantages that: (1) the attachment growth of Vero cells in a tissue culture vessel and on the surface of the micro-carrier can be supported, and the cells can be transferred from a serum culture medium to the serum-free culture medium without a course of adaptation; (2) the serum-free culture medium contains no animal origin components, and has basically definite chemical components, and a low cost; and (3) the cells grow well, and the cellular morphology, the density and the vitality are basically the same as those of the serum culture medium.

The invention discloses a culture method and serum-free culture medium composition of NK cells. The serum-free culture medium composition of the NK cells comprises an induction culture medium, a proliferation culture medium and an activation culture medium; the induction culture medium comprises a basal culture medium and an induction component, the proliferation culture medium comprises a basal culture medium and a proliferation culture component, and the activation culture medium comprises a basal culture medium and an activation component. The culture method of the NK cells comprises the step of performing cell culture by sequentially using the induction culture medium, the proliferation culture medium and the activation culture medium in stages. The serum-free culture medium composition of the NK cells comprises the induction culture medium, the proliferation culture medium and the activation culture medium which are designed for the different influence requirements of the induction, proliferation and activation periods of the NK cells, and the cultured NK cells are superior to those cultured through an existing technology on the aspects such as the total cell number, the proliferation speed, the amplification times and the killing activity on tumor cells.

the invention discloses a non-serum substratum for cell of HEK 293, which is composed of transferring, insulin, Primatone, lipoid compound, amino acid, inorganic salt and vitamin based on basic substratum.

The invention discloses a porous chitosan scaffold, and a neural stem cell porous chitosan scaffold and application thereof. A method for preparing the porous chitosan scaffold comprises the following steps of: dissolving chitosan in acetic acid to form 2 percent solution, adding the solution into pores of a cell culture plate, freeze-drying, adding NaOH into the pores of the cell culture plate for hydration, and freeze-drying twice at the temperature of -60 DEG C to obtain the porous chitosan scaffold. A method for preparing the neural stem cell porous chitosan scaffold comprises the following steps of: inoculating neural stem cell clone spheres to the sterilized porous chitosan scaffold in the culture spores, and adding a DMEM/F12 serum-free culture medium or an NGF-containing DMEM/F12 serum-free culture medium for culture to obtain the neural stem cell porous chitosan scaffold. The chitosan porous scaffold prepared by a low-temperature freeze-drying method has uniform pores, the porosity of 90 percent, the average pore size of 50 to 350mu m and high biocompatibility with neural stem cells (NSCs). Traumatic brain injury (TBI) is treated by transplanting the NSCs using chitoson as a carrier, and cognitive functions after injury, such as learning, memorizing and the like can be obviously improved.

The invention relates to a method for culturing an autologous peripheral blood lymphocyte. The method comprises the following steps: (1) separating a mononuclear cell from peripheral blood, resuspending in an X-VIVO15 serum-free culture medium to obtain cell concentration of 1*10<6>/mL, and culturing for 3 days; (2) supplementing the X-VIVO15 serum-free culture medium to 100 mL, adding IL-21*10<3> U/mL, and culturing for 1 day; (3) supplementing the X-VIVO15 serum-free culture medium to 200-240 mL, adding IL-21*10<3> U/mL, and culturing for 3 days; (4) supplementing the X-VIVO15 serum-free culture medium to 1000 mL, adding IL-21*10<3> U/mL, CTLA-4mAb100n g/mL and PD-1mAb100n g/mL; and (5) culturing for 5-7 days to prepare the autologous peripheral blood lymphocyte. The method disclosed by the invention can be used for improving the activation efficiency and amplification efficiency of an effector cell group by adding multiple monoclonal antibodies and cell factors to the X-VIVO15 serum-free culture medium, and can be used for effectively reducing the content of T regulatory cells by covering CTLA-4 and PD-1 molecules of the surfaces of all CIK cells by loading CTLA-4 and PD-1 antibodies in vitro especially, thus further enhancing the killing effect of the CIK cells on tumors.