37 results about "Monolayer culture" patented technology

The present invention provides a non-mouse, including human, pluripotential embryonic stem cell which can:(a) be maintained on feeder layers for at least 20 passages; and(b) give rise to embryoid bodies and multiple differentiated cell phenotypes in monolayer culture.The invention further provides a method of making a pluripotential embryonic stem cell comprising culturing germ cells and germ cell progenitors in a composition comprising a growth enhancing amount of basic fibroblast growth factor, leukemia inhibitory factor, membrane associated steel factor, and soluble steel factor to primordial germ cells under cell growth conditions, thereby making a pluripotential embryonic stem cell.Also provided are compositions useful to produce the pluripotent embryonic stem cells and methods of screening associated with the method of making the embryonic stem cell.

The invention provides culture platforms, cell media, and methods of differentiating pluriptent cells into hematopoietic cells. The invention further provides pluripotent stem cell-derived hematopoietic cells generated using the culture platforms and methods disclosed herein, which enable feed-free, monolayer culturing and in the absence of ER formation. Specifically, pluripotent stem cell-derived hematopoietic cell of this invention include, and not limited to, iHSC, definitive hemogenic endothelium, hematopoietic multipotent progenitors, T cell progenitors, NK cell progenitors, T cells, and NK cells.

The invention provides culture platforms, cell media, and methods of differentiating pluripotent cells into hematopoietic cells. The invention further provides pluripotent stem cell-derived hematopoietic cells generated using the culture platforms and methods disclosed herein, which enable feed-free, monolayer culturing and in the absence of EB formation. Specifically, pluripotent stem cell-derived hematopoietic cell of this invention include, and not limited to, iHSC, definitive hemogenic endothelium, hematopoietic multipotent progenitors, T cell progenitors, NK cell progenitors, T cells, NK cells, NKT cells and B cells.

The invention provides culture platforms, cell media, and methods of differentiating pluripotent cells into hematopoietic cells. The invention further provides pluripotent stem cell-derived hematopoietic cells generated using the culture platforms and methods disclosed herein, which enable feed-free, monolayer culturing and in the absence of EB formation. Specifically, pluripotent stem cell-derived hematopoietic cell of this invention include, and not limited to, iHSC, definitive hemogenic endothelium, hematopoietic multipotent progenitors, T cell progenitors, NK cell progenitors, T cells, and NK cells.

The invention provides culture platforms, cell media, and methods of differentiating pluripotent cells into hematopoietic cells. The invention further provides pluripotent stem cell-derived hematopoietic cells generated using the culture platforms and methods disclosed herein, which enable feed-free, monolayer culturing and in the absence of EB formation. Specifically, pluripotent stem cell-derived hematopoietic cell of this invention include, and not limited to, iHSC, definitive hemogenic endothelium, hematopoietic multipotent progenitors, T cell progenitors, NK cell progenitors, T cells, NK cells, NKT cells and B cells.

There is provided a method for the isolation and cultivation of mesenchymal stem / progenitor cells from umbilical cord blood, and also to a method for the differentiation of the umbilical cord blood-derived mesenchymal stem / progenitor cells into various mesenchymal tissues. The method include overlaying umbilical cord blood onto Ficoll-Hypaque solution; centrifuging the umbilical cord blood on the Ficoll-Hypaque solution to obtain a mononuclear cell layer; reacting cells obtained by monolayer culture of the mononuclear cells with antibodies to mesenchymal stem / progenitor cell-specific antigens for a predetermined period of incubation time; isolating only cells bound to their corresponding antibodies using a cell sorter; and cultivating the isolated cells, thereby obtaining mesenchymal stem / progenitor cells with high purity and excellent viability.

There is provided a method for growing human intervertebral cells. Disc tissue is surgically removed from a normal disc of a patient, the cells expanded by feeding with a cell stimulant such as a growth factor, or a cytokine or a bioactive agent to form monolayer primary cell cultures on a plastic mesh such as a nylon mesh. In the case of a growth factor, fetal bovine serum is preferred as it improves cell proliferation and production of appropriate extracellular matrix components. In another aspect of this invention, the monolayer primary cell cultures are seeded in alginate or agarose and fed again with the cell stimulant until three-dimensional cell cultures are formed. The cells are recovered from the alginate or agarose or from monolayer cultures. Re-implantation is carried out using bioresorbable carriers or cell suspensions.

The invention discloses a spraying type soilless culture vegetable bed and a culturing method thereof. The invention relates to an agricultural culture technology, in particular to a soilless vegetable culturing technology. The invention aims at solving the problems of energy conservation, consumption reduction and production and effect increase, and the like, in the process of soilless vegetable culturing in the reality. The spraying type soilless culture vegetable bed and the culturing method thereof can carry out both monolayer culture and multilayer culture and adopt a mechanical spraying and liquid feeding mode, the temperature can be adjusted by a solar water heater, the illumination can be adjusted by a fluorescent light and various situations in the vegetable growing process can be monitored by a computer.

The invention relates to a method for preparing and separating stereotyped endoderm cells by using monolayer culture technology and also discloses culture mediums suitable for preparing the stereotyped endoder mcells. The culture mediums comprise Neurobasal culture medium, DMEM / F12 culture medium, N-2 additive, B-27 additive, bovine serum albumin, mono-thioglycerin and activin A. The inventive method is simple and efficient, and can improve the efficiency in differentiating stereotyped endoderm of embryonic stem cells (preferably inhuman mammals) greatly.

The invention discloses an intestinal epithelium monolayer culture and characterization method based on mouse intestinal stem cells. The method comprises the following steps: (1) monolayer culture ofmouse intestinal epithelium; (2) characterization of monolayer culture of the intestinal epithelium. Compared with the prior art, the method is based on small intestinal crypts containing stem cells,optimizes the monolayer in-vitro culture system and culture medium of the intestinal epithelium, and includes a complete culture and characterization method.

The invention provides an in-vitro cartilage cell culturing method. The in-vitro cartilage cell culturing method comprises the following steps: acquiring a cartilage cell sample; performing primary culture on the cartilage cell sample; performing subculture on cartilage cells obtained after the primary culture, and when the convergence degree of the cartilage cells reaches 70-80% during the subculture, replacing a fresh culture medium and adding sulforaphane (SFN) to treat the cartilage cells. By the in-vitro cartilage cell culturing method, through the primary culture, morphs and specific molecular expression levels of the cartilage cells in the early stage are kept to be the same as those in a body to the maximum extent, so that morphologic characteristics of the cartilage cells in tight joint with a parent tissue can be displayed better; through the subculture, the cartilage cells are massively propagated, and at an appropriate time, the cartilage cells are degraded and inhibited by using the SFN, so that a degradation phenomenon during in-vitro monolayer culture can be effectively relieved, and a novel excellent method is provided for culturing the clinical cartilage cells.

There is provided a method for growing human intervertebral cells. Disc tissue is surgically removed from a normal disc of a patient, the cells expanded by feeding with a cell stimulant such as a growth factor, or a cytokine or a bioactive agent to form monolayer primary cell cultures on a plastic mesh such as a nylon mesh. In the case of a growth factor, fetal bovine serum is preferred as it improves cell proliferation and production of appropriate extracellular matrix components. In another aspect of this invention, the monolayer primary cell cultures are seeded in alginate or agarose and fed again with the cell stimulant until three-dimensional cell cultures are formed. The cells are recovered from the alginate or agarose or from monolayer cultures. Re-implantation is carried out using bioresorbable carriers or cell suspensions.

A 2D organoid for infection and culture of human diarrhea virus epithelial cell stem cell, which is obtained by Step 1, in which: human intestinal epithelial stem cells, human intestinal epithelial cells, or a tissue that includes at least one of these cells is subjected to three-dimensional culturing in the extracellular matrix; and a 3D organoid is obtained, and Step 2, in which: the 3D organoids obtained in Step 1 are dispersed to prepare single cells; the single cells are subjected to monolayer culturing in the extracellular matrix; and a 2D organoid is obtained in which the epithelial cells, which include differentiated trophoblastic cells and caliciform cell, and which constitute part of the human intestinal cavity, have a single-layer structure.

There is provided a method for growing human intervertebral cells. Disc tissue is surgically removed from a normal disc of a patient, the cells expanded by feeding with a cell stimulant such as a growth factor, or a cytokine or a bioactive agent to form monolayer primary cell cultures on a plastic mesh such as a nylon mesh. In the case of a growth factor, fetal bovine serum is preferred as it improves cell proliferation and production of appropriate extracellular matrix components. In another aspect of this invention, the monolayer primary cell cultures are seeded in alginate or agarose and fed again with the cell stimulant until three-dimensional cell cultures are formed. The cells are recovered from the alginate or agarose or from monolayer cultures. Re-implantation is carried out using bioresorbable carriers or cell suspensions.

This invention relates to a composition comprising a synergistic combination of ketoconazole and sulforaphane for use in skin lightening. This invention is based on the fact that a combination of ketoconazole and sulforaphane has been found to synergistically inhibit pigment production in B16 monolayer cultures. Thus the composition, when applied topically or imbibed over an appropriate length of time in-vivo, would be expected to cause skin lightening, or to reduce blemishes and / or hyperpigmented spots and / or solar lentigines leading to an improvement in skin tone and evenness.

The invention belongs to the technical field of cell culture loading materials, and particularly relates to a preparation method of a modified silicon wafer loading material and application of the modified silicon wafer loading material in cell culture. According to the invention, gelatin macromolecules are loaded on the surface of a polished silicon wafer, and then a gelatin coating with micropores is formed on the surface of the silicon wafer by using a lyotropic phase separation method or a gelatin coating with a specific three-dimensional topological structure is prepared by using a micro-pattern printing technology; and thus, the modified silicon wafer cell loading material with a micropore or fiber topological structure on the surface is prepared. The modified silicon wafer overcomes the limitation of the hydrophobicity of a silicon wafer material and a single-layer culture mode, not only has good biocompatibility, but also is more beneficial to the adhesion and growth of cells due to the three-dimensional gelatin coating structure of the modified silicon wafer, and can effectively simulate the three-dimensional environment of cell growth; meanwhile, under the limited volume, a larger loading area can be provided, and the modified silicon wafer loading material is suitable for being used as a large-scale cell culture material.

The invention provides a human chondrocyte culture medium, and belongs to the technical field of cells. The invention optimizes a traditional culture medium, and by adding exosomes, fetalbovine serumand penicillin-streptomycin into a basal culture medium, chondrocyte proliferation can be effectively promoted, dedifferentiation of human chondrocytes is prevented, transformation of the human chondrocytes into fibrocyte-like phenotypes is prevented, and the single-layer culture passage number is increased. The invention also provides a method for isolating culture of the human chondrocytes by using the above-mentioned culture medium, and normal chondrocytes cultured to the sixth passage can be obtained.

The present invention relates to a differentiation inducing method comprising: introducing, into human induced pluripotent stem cells, a combination of genes of transcription factors which are possessed by fetal and adult hepatic cells, but are not possessed by human induced pluripotent stem cells; and adhering the human induced pluripotent stem cells to a substrate for monolayer culture in a medium containing a combination of growth-promoting agents for proliferation and differentiation, thereby enabling induction of the differentiation of the human induced pluripotent stem cells in large amounts in a short term, a hepatic progenitor cell produced by the method, and a cell composition for transplantation into the liver comprising the cell. The transcription factors include FOXA2, GATA4, HEX and C / EBPα, and the growth-promoting agents include oncostatin M, an epidermal growth factor, retinoic acid, dexamethasone, insulin, transferrin and a selenite ion.

The invention discloses a preparation method of a human 3D structure epidermal cell sheet and a clinical application thereof. The preparation method is characterized by comprising the following steps that (1), human epidermal stem cells, base layer horn cells, melanocyte stem cells and the like are separated at a time through a Wbbase mixed enzyme; (2), two-step culture is carried out, wherein the two-step culture comprises a) non-serum mixing single-layer culture of three kinds of cells and b) induced differentiation of cells and culture of the 3D structure epidermal cell sheet; (3), the cell sheet is stripped; and (4), the cell sheet is clinically applied to treat leucoderma, burn and other skin pigment depigmentation diseases. The creativity of the preparation method of the human 3D structure epidermal cell sheet and the clinical application thereof is that three kinds of cells are separated simultaneously, the effective time is prolonged by 150-300 times compared with that of traditional trypsin separation methods, activity of the enzyme does not need to be neutralized, the activity of separated cells is greater than 95%, and the total yield is greater than 1-5*107 / g of skin; the two-step culture method amplifies the three kinds of cells at the same time, so that the amplification fold is improved by five times; the melanin forming unit is close to a normal person; and the compound color effect of transplanted skin is close to or consistent with the skin of a person.

The invention discloses a culture method for increasing the virus titer of duck flavivirus, and belongs to the field of microbial virus technologies. The culture method mainly comprises the following steps: preparing a DMEM culture solution; after fetal calf serums are added, inoculating cells for carrying out monolayer culture; after an affinity absorber and the DMEM culture solution are added, further culturing for 1-30 min, and inoculating duck flavivirus for 2-5 days, so that duck flavivirus liquid with a high virus titer can be obtained. The culture method disclosed by the invention increases the contact affinity of viruses and cells mainly by changing a microenvironment between the duck flavivirus and the cells, thereby greatly increasing the virus titer of the duck flavivirus.

The invention relates to a method for preparing and separating stereotyped endoderm cells by using monolayer culture technology and also discloses culture mediums suitable for preparing the stereotyped endoder mcells. The culture mediums comprise Neurobasal culture medium, DMEM / F12 culture medium, N-2 additive, B-27 additive, bovine serum albumin, mono-thioglycerin and activin A. The inventive method is simple and efficient, and can improve the efficiency in differentiating stereotyped endoderm of embryonic stem cells (preferably inhuman mammals) greatly.

The present invention relates to monolayer cardiac differentiation techniques utilizing defined conditions providing feeder-free monolayer culture systems, serum-based or serum free, and applicable to both healthy control and patient derived stem cells.

The invention discloses an in-vitro monolayer culture and representing method for constructing mouse intestinal epithelium by using Transwell. The method comprises the following steps of (1) respectively inoculating intestinal tract crypt structures containing intestinal tract stem cells and intestinal tract epithelial myofibroblasts to the upper surface and the lower surface of a Transwell nestedpolycarbonate membrane, nesting Transwell in a porous cell culture plate, adding an upper-layer cell and a lower-layer cell into an optimized culture medium, and carrying out co-culture to form a single-layer intestinal tract epithelium in vitro; and (2) regularly detecting the transepithelial resistance value of the single-layer intestinal epithelium every day in the culture process, representing that the intestinal in-vitro single-layer culture and the in-vivo intestinal epithelium have the same cell composition and structural characteristics by immunofluorescence and electron microscope results, and analyzing the permeability of the single-layer intestinal epithelium by taking isothiocyanate labeled glucan as a tracing reagent. The intestinal tract crypt and intestinal tract epithelialmyofibroblasts are co-cultured for the first time to form an in-vitro intestinal tract epithelial monolayer culture model with similar structural characteristics, cell composition and permeability toin-vivo intestinal tract epithelium.

The present invention relates to regulatory cells, which are capable of restoring, maintaining or improving the stable cartilage phenotype of expanded and passaged chondrocytes. These regulatory cells are also capable of directing precursor and stem cells into the chondrogenic lineage. An enriched population of regulatory cells can be obtained by harvesting the non-adherent cells in the culture medium of a monolayer culture of PO chondrocytes.

The invention discloses a preparation method of a gardnerella vaginalis culture medium, comprising the steps of: step 1, accurately weighing 4.1g of Columbia blood agar base, and adding 95ml of distilled water for dissolving with heating; step 2: autoclaving the dissolved solution of the step 1; step 3: cooling the dissolved solution to about 50 DEG C, adding 5ml of defibered rabbit blood and combined bacteriostatic agent under the condition of sterile operation; step 4, pouring the solution into a sterilized glass plate, regarding the solution as a rabbit blood Columbia agar monolayer culture medium, cooling the solution for later use; step 5: inoculating the prepared rabbit blood Columbia agar monolayer culture medium with gardnerella vaginalis, carrying out partitioned streak inoculation; and step 6: carrying out incubation at 35 DEG C with 5% CO2 for 48 hours. The preparation method only uses two components namely the rabbit blood and Colombia blood agar as nutrient substances to prepare the monolayer culture medium, can achieve the effect of separating the gardnerella vaginalis, realizes the characteristics of low cost, convenient operation, easy availability of raw materials and the like in preparation of culture media, and is favorable for wide development in different levels of hospitals.

The neural cell population of the present invention is characterized in that in the presence of F and FGF-2, the cells are in an adherent monolayer culture, and at least 80% of the cells are symmetrically divided neural stem cells, No more than 1% of the cells express markers of mature astrocytes, neurons, or oligodendrocytes, and they express the markers RC2, 3CB2, and BLBP, and neural stem cells in said population are also characterized in that they express At least one of the proteins: GLAST, Pax‑6, neural progenitor cell markers stem protein or vimentin, LewisX antigen, Musashi‑l or convexin. The preparation method of the invention is simple and easy to operate.

A 2D organoid for infection and growth culture of human diarrhea virus, obtained by: (1) obtaining a 3D organoid by three-dimensionally culturing a human intestinal epithelial stem cell, a human intestinal epithelial cell, or a tissue containing at least any of those cells on an extracellular matrix; and (2) dispersing the 3D organoid to prepare a single cell, and monolayer-culturing the single cell on an extracellular matrix to obtain the 2D organoid having a monolayer structure in which epithelial cells contain differentiated trophoblastic cells and goblet cells and configured as human intestinal lumen having a monolayer structure.