30 results about "Feeder Layer" patented technology

The invention provides cell culture conditions for culturing stem cells, including feeder-free conditions for generating and culturing human induced pluripotent stem cells (iPSCs). More particularly, the invention provides a culture platform that allows long-term culture of pluripotent cells in a feeder-free environment; reprogramming of cells in a feeder-free environment; single-cell dissociation of pluripotent cells; cell sorting of pluripotent cells; maintenance of an undifferentiated status; improved efficiency of reprogramming; and generation of a naïve pluripotent cell.

The invention provides a method for culturing primary spermatogonial stem cells of Tibetan sheep, and belongs to the technical field of cell separation and culture. The culture method provided by the invention comprises the following steps: disinfecting and cleaning testis tissues of Tibetan sheep lambs; shearing tissue blocks, and digesting and cleaning to obtain a single-cell suspension of the seminiferous tubule; purifying the single-cell suspension, and culturing without a feeding layer or with the feeding layer; the culture without the feeding layer refers to culture in a laminin-coated culture container by using a feeding-layer-free MEM alpha culture medium or a StemProTM-34SFM complete culture medium; culture with a feeding layer refers to culture in a mouse fibroblast line feeding layer or a Tibetan sheep primary testicular sertoli feeding layer, and a feeding layer MEMS alpha culture medium is added. According to the culture method provided by the invention, the Tibetan sheep spermatogonial stem cells with higher purity can be obtained through separation, the subsequent proliferation of the spermatogonial stem cells can be remarkably promoted, and the apoptosis of the spermatogonial stem cells can be inhibited.

Disclosed is a chemically defined airway basal stem cell culture medium. The provided culture medium involves a simple preparation method and high safety, and does not require a feeder layer or a matrigel-coated culture plate. By means of the culture medium, primary cultured airway basal stem cells achieve good stability; and after multiple passages, the normal morphology of the basal stem cells can be maintained and good differentiation potential can be maintained. In addition, the culture medium has clear components, and can meet both scientific research-grade and clinical-grade applications of airway basal stem cells.

The invention belongs to the technical field of cell biology, and particularly relates to a feeder-layer-free and serum-free bovine embryonic stem cell culture medium, a culture system and a culture method. The bovine embryonic stem cell culture medium comprises a conditioned culture medium and additives, the additives are IWR-1 and activin A, and the concentrations of the IWR-1 and the activin A in the conditioned culture medium are 2.5-5 [mu] M and 20-25 ng / mL respectively; the culture system comprises a bovine embryonic stem cell culture medium, gelatin with the mass concentration of 0.1-0.2% and a subculture medium, and the gelatin with the mass concentration of 0.1-0.2% is used for providing an attachment matrix for bovine embryonic stem cells; the subculture medium is prepared by adding Y27632 into a bovine embryonic stem cell culture medium. The cost of the extracellular matrix is reduced, and the feeder-layer-free bovine embryonic stem cell has high single cell passage efficiency and is convenient for gene editing operation.

The invention provides a method for producing functional spinal cord GABAergic neural precursor cells from human pluripotent stem cells, which successfully realizes directional differentiation of the human pluripotent stem cells by adopting a feeder-layer-free culture method based on a three-stage in-vitro induced differentiation strategy. The invention further provides application of the cells obtained through the method in treatment of spinal cord injury, a novel strategy based on the functional spinal cord GABAergic neural precursor cells is provided, a new thought and means are provided for treatment of spinal cord injury, and stem cell treatment is expected to proceed towards clinical precision and industrial application.

This invention discloses a method for preparing NK cells, the NK cells obtained thereby, and their applications. The method includes: sorting NK cells from peripheral blood of an allogeneic donor; culturing them in a serum-free medium containing CD3 monoclonal antibody, CD16 antibody, and IL-2, without the need for a feeder layer; preferably, IL-21 is added for amplification on days 3-5 of culture, and the cells are cultured for 20-40 days. This method yields non-genetically modified NK cells, among which CD56... bright The NK cells comprise >90% of the cell line and highly express a variety of activating and functional molecules. These NK cells effectively clear Aβ oligomers and senescent cells, and alleviate neuroinflammation. In treatment, intracranial injection achieves equivalent or better efficacy with only about 1 / 10th the dose administered intravenously, significantly clearing Aβ deposits in the brains of AD model mice and improving pathology.

The present invention provides cell culture conditions for culturing stem cells, including feeder-free conditions for producing and culturing human induced pluripotent stem cells (iPSCs). More specifically, the present invention provides a culture platform that allows long-term culture of pluripotent cells in a feeder-free environment; reprogramming of cells in a feeder-free environment; single cell dissociation of pluripotent cells; cell sorting of pluripotent cells; maintenance of an undifferentiated state; increased efficiency of reprogramming; and the generation of original pluripotent cells.

This invention relates to the field of bovine embryonic stem cell culture technology, and particularly to a long-term culture system for morphological bovine embryonic stem cells completely without a feeder layer and its applications. After trying various culture methods, this invention establishes a feeder-free bovine morphological embryonic stem cell line (FF-bESCs). Bovine morphological embryonic stem cells cultured under the feeder-free conditions provided by this invention can maintain stable proliferation capacity and pluripotency over a long period. The bovine morphological embryonic stem cells provided by this invention can undergo gene editing and can be effectively induced to differentiate into primordial germ cell-like cells (PGCLCs). This invention lays an important foundation for the standardized and large-scale culture of bovine embryonic stem cells under feeder-free conditions, and for their application in stem cell breeding and other fields.

The application belongs to the technical field of cell biology, and particularly relates to a feeder-free and serum-free bovine embryonic stem cell culture medium, a culture system and a culture method. The bovine embryonic stem cell culture medium comprises a conditioned medium and an additive. The additive is IWR-1 and activin A. The concentrations of IWR-1 and activin A in the conditioned medium are 2.5-5 muM and 20 ng / mL-25 ng / mL respectively. The culture system comprises the bovine embryonic stem cell culture medium, 0.1%-0.2% gelatin in mass concentration and a subculture medium. The 0.1%-0.2% gelatin in mass concentration is used to provide an attachment matrix for the bovine embryonic stem cell. The subculture medium is prepared by adding Y27632 to the bovine embryonic stem cell culture medium. The application reduces the cost of extracellular matrix, and the feeder-free bovine embryonic stem cell has high single cell subculture efficiency and is convenient for gene editing operation.

The present disclosure provides feeder hydrogel particles that can function to support the growth, proliferation, and / or activation of a target cell in culture. The present disclosure also provides methods of culturing target cells with feeder hydrogel particles.

The invention discloses a human airway basal stem cell culture medium as well as a preparation method and application thereof. The culture medium provided by the invention is simple in preparation method and high in safety, a culture plate does not need to be coated with a feeding layer or matrigel, and the airway basal stem cells subjected to primary culture are good in stability; after multiple passage, the normal form of the basal stem cells can be maintained, and good differentiation potential can be maintained; meanwhile, components of the culture medium are clear, and scientific research-grade and clinical-grade application of the airway basal stem cells can be met at the same time.

To provide a defined culture medium for isolating stratified epithelial stem cells and stably maintaining the epigenetics of the stratified epithelial stem cells during multiple passages, and to provide clonal stratified epithelial stem cells isolated using the defined culture medium.SOLUTION: And a defined culture medium comprising each of ROCK (Rho kinase) inhibitors, mitogenic growth factors, insulin or IGF, TrkA inhibitors, and Oct4 activators, and comprising at least one of VEGF inhibitors, tyrosine kinase inhibitors, and / or FGF10 or FGF10 agonists, and optionally further comprising TGF β signaling pathway inhibitors and / or bone morphogenetic proteins (BMP) antagonists, wherein the defined culture medium supports epigenetically stable growth and proliferation of stem cells of stratified epithelial tissue origin in culture.SELECTED DRAWING: Figure 1

The invention provides a keratin 15 type hair follicle stem cell culture medium, an adherent culture method and application, and belongs to the technical field of stem cell culture. The invention provides a keratin 15 type hair follicle stem cell culture medium, which comprises a basic culture medium, a serum substitute and a nutrition additive, a mixed culture medium containing a DMEM basic culture medium and an F12 basic culture medium is used as the basic culture medium, and an epidermal growth factor, a platelet lysate and a bovine pituitary extract are used as the serum substitute. The keratin 15 type hair follicle stem cells are cultured on the basis of the culture medium, proliferation-terminated autologous mesenchymal cells serve as a feeding layer, the activity of the hair follicle stem cells p63 can be reserved, and the keratin 15 positive hair follicle stem cells are amplified.

A method of differentiating nonhuman primate induced pluripotent stem cells (iPSCs) to oligodendrocytes is provided. The method may include providing a modified vector comprising a piggyBac vector backbone that expresses one or more differentiation factor genes, including SOX10, OLIG2, and NKX6-2. The piggyBac vector backbone may comprise one or more terminal inverted repeats and one or more transposase recognition sites configured to interact with a piggyBac transposase to mediate genomic integration at TTAA or noncanonical target sequences. The vector may further include a selectable marker and a promoter for regulated expression. The modified vector may be introduced into nonhuman primate iPSCs, for example from Macaca fascicularis, by electroporation and used in a feeder-free differentiation workflow. The resulting differentiated cells may be identified by expression of oligodendrocyte-associated markers including O4, O1, PDGFRA, MOG, GALC, CNPase, and myelin basic protein (MBP).

To provide a novel method that enables culture in a suspension state in establishment and proliferation of stem cells.SOLUTION: A method for producing pluripotent stem cells, comprising a step of culturing pluripotent stem cells under feeder-free and suspension culture conditions in a medium containing a scaffold material including atelocollagen and separated from a culture vessel.SELECTED DRAWING: None

The invention provides an in vitro method for producing an isolated mammalian primitive retina stem cell (pRSC), comprising: (a) culturing an isolated embryonic stem cell (ESC) from a mammal in a cell culture medium that is free of feeder cells, feeder layer conditioned medium, or serum to produce and grow a culture of the isolated ESC; and (b) contacting the culture of the isolated ESC so grown with one or more of a Wnt or TGF-beta / BMP signaling inhibitor to differentiate the isolated ESC of (a) into a primitive retina stem cell, thereby producing an isolated mammalian pRSC.

The invention provides a method for inducing efficient amplification of NK cells through the synergistic effect of exosome and immunosuppressor, and belongs to the technical field of cell culture, and the method comprises the following steps: 1) extracting and purifying stem cell exosome; the stem cells are human umbilical cord mesenchymal stem cells; 2) separating CBMC cells and plasma from the umbilical cord blood, inactivating the plasma, activating platelets, incubating and separating to obtain super plasma; 3, CBMC cells are inoculated and cultured in a low-oxygen incubator for 10-16 h, then the exosomes in the step 1 are added, culture continues to be conducted for 10-16 h, the CBMC cells are transferred to a normal oxygen environment, culture does not exceed 4 h, a culture medium and super plasma are supplemented, culture continues to be conducted, and amplified NK cells are obtained.According to the technical scheme, in-vitro culture of the high-activity NK cells can be efficiently amplified and maintained without a feeding layer is achieved.

The present invention relates to a culture media system that useful for the isolation and epigenetically stable propagation of normal stem cells in culture which are derived from columnar epithelial tissues and cancer stem cells from epithelial cancers. In certain embodiments, the culture system is a feeder-free system.

The invention discloses application of tetracycline drugs in synchronous separation and maintenance of bovine embryonic stem cells and bovine trophoblast stem cells. Belongs to the technical field of cytobiology and animal embryo engineering. According to the method, cattle blastocysts or cell clusters in the cattle blastocysts are cultured in a culture medium containing tetracycline drugs, so that the dryness of bESCs and bTSCs is synchronously separated and maintained in the same culture system. The technical problem of synchronous separation of bESCs and bTSCs is solved by utilizing the bidirectional regulation and control capability of tetracycline drugs for the first time, and an efficient and low-cost standardized platform is provided for artificial embryos, regenerative medicine and agricultural bioengineering.

An mRNA-LNP-based reprogramming method for induced pluripotent stem cells. Specifically, a cationic lipid is used to encapsulate an mRNA mixture of a plurality of reprogramming factors to prepare a nano-liposome containing the reprogramming factors; a certain amount of the nano-liposome containing the reprogramming factors is dropwise added into adult cells in a feeder-free system to obtain reprogrammed induced pluripotent stem cells, thereby achieving a low-cost, low-toxicity, efficient, stable, repeatable, and safer iPSCs reprogramming technology.

The present application relates to a method of deriving a human embryonic stem cell (hESC) line under feeder-free and heterologous conditions.

The invention relates to the technical field of biotechnology, and discloses a construction method of a bovine originating embryonic stem cell line, which comprises the following steps: (1) obtaining in vitro fertilized bovine early blastocyst; (2) trophoblast cells of the blastocyst are removed, an inner cell cluster is inoculated into a culture system containing a feeder layer, and the culture system is a basic culture medium added with FGF2 and WNT pathway inhibitors; and (3) culturing until an inner cell cluster forms a cell set drop, and carrying out digestion passage to obtain the stably proliferated bovine originated embryonic stem cell line. According to the construction method and the application of the bovine embryonic stem cell line, a specific initial induction combination of 'FGF2 + WNT inhibitor IWR1' is adopted, and a cell mass in an early blastocyst is used for establishing the line, so that the bovine embryonic stem cell line which is uniform in form, can be stably passaged for more than 26 generations and still keeps typical embryonic characteristics is successfully obtained; the problems of unstable state and easy differentiation of the bovine stem cell line in the prior art are solved.

The present disclosure relates generally to novel methods and compositions for using engineered reprogramming factor(s) for the creation of induced pluripotent stem cells (iPSCs) through a kinetically controlled process. Specifically, this disclosure relates to establishing combinations of reprogramming factors, including fusions between conventional reprogramming factors with transactivation domains, optimized for reprogramming various types of cells. More specifically, the exemplary methods disclosed herein can be used for creating induced pluripotent stem cells from various mammalian cell types, including human fibroblasts. Exemplary methods of feeder-free derivation of human induced pluripotent stem cells using synthetic messenger RNA are also disclosed.

The present application relates to the field of cell biology, specifically will be by adding phosphoantigen (HMBPP), IL-2 and other factors, specific activation of gamma delta T cells, and Daudi lymphoma cell feeder layer co-culture, promote the expansion of gamma delta T cells, at the same time, promote the memory phenotype and survival of gamma delta T cells, and improve the cytotoxicity and function of gamma delta T cells. The method simultaneously induces peripheral blood mononuclear cells to become gamma delta T cells and activates gamma delta T cells, enhances immune signal transduction, and significantly improves the activation efficiency and expansion capacity. The results show that the proliferation of gamma delta T cells is 742.62 times in 14 days, which is significantly higher than that of conventional CIK culture; CD3 + cell ratio reaches 90.15%, which is better than 75.29% of CIK. Thus, a short cycle, high yield, and good purity gamma delta T cell preparation process is formed, which embodies the creativity of multi-factor synergy and feeder layer co-culture, and brings significant beneficial effects.

Cord blood or peripheral blood NK cells are prepared from whole blood mononuclear cells without the need to isolate CD34+ hematopoietic stem cells or NK cells, and without the need for a feeder layer. Advantageously, the methods presented herein use an enrichment process that uses antiCD16 agonist antibodies, antiCD3 antibodies, and N-803. Moreover, contemplated processes are suitable for adaptation into a fully automated production process (GMP in a box).

The present invention aims to provide a method for producing iPS cell-derived keratinocytes that can induce keratinocytes that highly express keratinocyte markers in a feeder-free and serum-free environment. [Solution] A method for producing iPS cell-derived keratinocytes, comprising an induction step of inducing iPS cells into keratinocytes using one or more induction media in a feeder-free and serum-free environment, wherein for at least a portion of the induction step, a medium containing fibroblast growth factor 7 (FGF7) is used as the induction medium.

The invention relates to the technical field of biology, in particular to a combined culture medium of chicken embryo primordial germ cells (Primordial Germ Cells, PGCs) and a cell culture method of the combined culture medium, calcium ion-free DMEM is used as a basis, various supplements, nucleoside, sodium pyruvate, amino acid, vitamins, additives, trace components, three antibodies, serum and specific cell factors are added, filtration sterilization is performed after osmotic pressure is adjusted, and the combined culture medium and the cell culture method of the combined culture medium are used for culturing the chicken embryo primordial germ cells (Primordial Germ Cells). The cell factors are added before use to adapt to the culture of the related PGCs. The method does not need a feeding layer, simplifies the operation, avoids exogenous pollution, improves the purity and survival state of the PGCs, promotes proliferation, has no sex preference, is adaptive to different sources of PGCs, is stable and reliable, and has a wide application prospect.

The invention relates to the technical field of biomedicine, in particular to a differentiation method of NK cells. By optimizing a differentiation strategy, culture medium components and culture parameters, the NK cell differentiation method of the induction system without the feeder layer is obtained; the differentiation method of the NK cells is low in cost and high in differentiation efficiency, and the obtained NK cells have a high cell killing effect.

To provide methods and compositions that allow stem cells to differentiate into secondary hematopoiesis without relying on co-culture or serum-containing medium and without requiring formation of an embryoid aggregate as an intermediate.SOLUTION: The present invention provides a method and composition for generating hematopoietic lineage cells via secondary hematopoietic endothelium (HE) and secondary hematopoietic stem cells (HSC) derived from pluripotent stem cells containing hiPSC by a scalable monolayer culture platform without requiring EB formation under serum / feeder free conditions. The present invention is also a method and composition for differentiating and expanding hematopoietic line cells from pluripotent stem cells in monolayer culture, comprising contacting pluripotent stem cells with a BMP pathway activator, optionally with bFGF.SELECTED DRAWING: None