202 results about "Matrigel" patented technology

The invention relates to an organ-like core-shell microsphere as well as a preparation method and application thereof. The preparation method comprises the steps that 1, raw materials are prepared, specifically, shell raw materials and inner core raw materials are prepared, the shell raw materials comprise a photoinitiator and methacrylated hyaluronic acid (HAMA), and the inner core raw materials comprise organoid precursor cell suspension and matrigel; 2) molding: extruding the dispersion phase of the shell raw material wrapping the core raw material to a continuous phase by using a micro-fluidic chip and a high-precision injection pump, cutting the dispersion phase into liquid drops by the continuous phase, and performing illumination curing on the liquid drops to form core-shell microspheres; and (3) incubating, namely culturing the core-shell microspheres in a culture medium, so that the organoid precursor cells in the inner core are developed into organoids. According to the preparation method, rapid forming and curing of the organ-like precursor cells carried by the matrigel are achieved, so that the time of the organ-like precursor cells staying in the oil phase is shortened, the cells can obtain oxygen and nutrient substances, and the cell activity is improved.

The invention discloses a construction method of a homogenized self-assembled heart organoid, and belongs to the field of organoid. The preparation scheme of the heart organoid provided by the invention is controllable in method and simple in procedure, emphasizes the transition of differentiation from an iPSC 2D level to a 3D self-assembly heart organoid, overcomes the heterogeneity, including size, structure, function and even gene difference, of cross-batch or even same-batch organoid caused by an existing differentiation method, and has good cross-batch reproducibility. The constructed heart organoid contains various cell types and can continuously beat for at least 180 days in vitro, and the heart organoid in the same batch is uniform in height and stable in function. The preparation scheme does not need an engineering scaffold, matrigel and the like, emphasizes self-assembly of heart organs, and better accords with physiological characteristics of a human body.

The invention provides a micro-fluidic chip capable of culturing oral squamous cell carcinoma organoid and a preparation method thereof, and relates to the technical field of micro-fluidic chips, the chip comprises a four-layer structure: the first layer is a closed layer, the second layer is a culture solution / drug pool layer (containing a concentration gradient injection channel and a slow flow channel), the third layer is a matrix culture pool layer, and the fourth layer is a matrix culture pool layer. The fourth layer is a matrigel inflow layer (containing an S-shaped flow channel); the second layer and the third layer are integrally formed, and all the layers are connected through oxygen plasma welding; the problems that in the prior art, matrigel is prone to shifting, drug gradient regulation and control is poor, operation is tedious, and adaptability is insufficient are solved, the matrigel and organoids can be stably fixed, the drug concentration gradient can be precisely regulated and controlled, the nutrition supply efficiency is enhanced, the structural integration level and operation convenience are improved, and the application prospect is wide. The method is suitable for oral squamous cell carcinoma organoid culture, drug screening and tumor mechanism research.

The invention provides preparation, sorting and application of LGE / CGE organoid and GABAergic neurons, and belongs to the field of brain organoid differentiation and GABAergic neurons. The preparation method of the LGE / CGE type organ comprises the following steps: S1, performing embryoid formation and initial culture on the 0th-4th day; s2, performing nerve induction and matrigel embedding on the 5th-11th day; s3, culturing the brain organoid on the 12th day to the 19th day; s4, dynamically culturing and screening the brain organoid on the 20th-35th day; and S5, carrying out differentiation and collection on LCOs of the LGE / CGE type organs after the 35th day. According to the method, the LGE / CGE organs containing the high-purity GABA energy neurons are obtained through differentiation from the brain organs for the first time, the high-purity LGE type GABA energy neurons and the high-purity CGE type GABA energy neurons are obtained through a specific sorting method, and the method has the deep application potential in the fields of disease model construction, medicine research and development and the like.

The invention belongs to the technical field of analysis and detection, and particularly relates to a beta-galactosidase response type hemicyanine near-infrared fluorescent probe, a preparation method thereof and application of the beta-galactosidase response type hemicyanine near-infrared fluorescent probe as a tumor organ imaging probe. The near-infrared fluorescent probe provided by the invention takes a water-soluble hemicyanine skeleton as a fluorophore, so that the biocompatibility of the near-infrared fluorescent probe is improved; based on the principle that beta-galactose can be selectively cut by beta-galactosidase, a galactose structure with a self-leaving group is used as a response group, selective quick response to the beta-galactosidase is achieved, and high sensitivity to the beta-galactosidase is shown. The near-infrared fluorescent probe provided by the invention can emit near-infrared fluorescence with long wavelength, has lower background fluorescence and good penetration depth, and can be directly used for detecting organoids containing the matrigel without eliminating the matrigel, so that the organoids are prevented from being damaged or additional experimental variables are prevented from being introduced in the process of eliminating the matrigel.

Provided are chemically modified silk fibroin and a use thereof in cell and organoid culture. The chemically modified silk fibroin is a product obtained by sequentially carboxylating silk fibroin and modifying same by an organic amine having a phenol group. Also provided is a hydrogel based on the chemically modified silk fibroin. A hydrogel system has a hierarchical structure and mechanical properties similar to those of an extracellular matrix, and exhibits characteristics such as definite composition, controllable physicochemical properties, low cytotoxicity, good biocompatibility, and biodegradability, can support the growth and differentiation of cells and organoids, and is suitable as a matrigel for cell and organoid culture.

The invention provides a mouse middle ear epithelium 3D organoid and a construction method thereof, and is characterized in that the method comprises the following steps: a, taking a bubble structure of a mouse middle ear tissue, cleaning, digesting, filtering, centrifuging filtrate, collecting precipitate, and separating primary cells of the mouse middle ear; and b, inoculating the digested single cells and cell agglomerates into a 24-pore plate, wrapping with matrigel, and carrying out 3D culture, wherein a medium is a middle ear epithelium amplification medium. And culturing for about 3-5 days to obtain the mouse 3D middle ear proliferation type organoid. And performing differentiation culture by using a middle ear epithelium differentiation medium to obtain the mouse middle ear epithelium organ. According to the method, the 3D mouse middle ear epithelium organ is successfully constructed, the physiological and pathological processes of the middle ear are conveniently analyzed and researched, the interaction between the middle ear microenvironment and cells can be more accurately simulated, and the application range is wide as a model.

The invention discloses a culture system, culture method and application of gastrointestinal tumor chemotherapy drug-resistant organoid, the culture system comprises: a basic culture medium, which is a DMEM / F12 culture medium, and is additionally added with a special growth promoting factor obtained from a natural extract; the multiple growth factors comprise an epidermal growth factor (EGF), a fibroblast growth factor (FGF), a bone morphogenetic protein (BMP) and a vascular endothelial growth factor receptor inhibitor (VEGFRi), and the concentration of each growth factor is accurately screened and optimized; the extracellular matrix is Matrigel modified by physical and chemical methods; the specific additive comprises an antioxidant N-acetylcysteine (NAC). According to the culture system, the culture method and the application of the gastrointestinal tumor chemotherapy drug-resistant organoid, the culture system which is closer to a real environment in vivo is constructed by optimizing basic culture medium components, setting various growth factors and specific additive concentrations and adopting modified Matrigel as an extracellular matrix.

The invention discloses a preparation method and application of a bovine tongue mucous membrane organoid, and relates to the technical field of organoid separation and culture. The preparation method of the ox tongue mucosa organoid comprises the following steps: treating fetal ox tongue tip tissues with digestive enzymes Dispase I, Collagenase II and Collagenase IV, separating ox tongue mucosa, cutting into pieces, treating again with pancreatin, and sieving to remove tissues and impurities; and continuously culturing by adopting a three-dimensional culture technology-matrigel embedding method and an organoid culture medium to obtain the final ox tongue mucous membrane organoid. The ox tongue mucous membrane organoid prepared by the preparation method disclosed by the invention has normal cell proliferation, differentiation and metabolism functions, and can be stably passaged under an in-vitro culture condition; the method can be widely applied to various related scientific researches such as FMDV infection mechanism research, drug screening and vaccine evaluation.

The invention relates to the technical field of biology, and particularly discloses a construction method and application of an endometrial epithelium organ fibrosis model. According to the method, human endometrial epithelial cells are extracted, and an endometrial epithelial organ is established by adopting a Matrigel three-dimensional culture system. Afterwards, the organoid is induced to form endometrial phenotypes in different physiological cycles through hormone stimulation, and a ferroptosis inducer FIN56 is added in the culture process to induce fibrosis. The expression of GPX4, Fibronectin and p-Smad2 / 3 is detected through Western blot and immunofluorescence, and the successful construction of the fibrosis model is verified. The invention provides an in-vitro model capable of simulating endometrial fibrosis pathological characteristics of a patient with PCOS (polycystic ovarian syndrome), the in-vitro model can be used for mechanism research and drug screening of PCOS-related endometrial lesions, and a new research tool and an intervention strategy are provided for treatment of PCOS endometrial fibrosis.

The invention relates to the technical field of organs, and discloses construction of a matrigel-free kidney organoid and a method for evaluating drug safety by using liver and gall-kidney organoid. By utilizing the culture medium set provided by the invention, the kidney organoid with more complete functions and structures can be constructed through iPSC induction, and matrigel does not need to be adopted in the construction and culture process, so that the construction operation of the organoid is greatly simplified, and the cost is saved. The kidney-like organ constructed by the invention has a basic structure of the kidney, has corresponding response to medicine components which are known in the field and have influence on the kidney, and can be used as a kidney in-vitro model.

The invention discloses a bladder cancer lymph node metastasis derived tumor organoid culture medium, a culture method and a rapid drug detection method, and belongs to the technical field of organoid and cell culture. The method comprises the following steps: culturing a bladder cancer lymph node metastasis-derived tumor organoid by using a bladder cancer lymph node metastasis-derived tumor organoid culture medium, immersing matrigel drops in the culture medium, and obtaining the bladder cancer lymph node metastasis-derived tumor organoid after the cultured organoid grows to 100-150 microns, to-be-detected drugs with different concentrations are directly added into the cell pore plate inoculated with the tumor organoids, the influence of the to-be-detected drugs on cell growth or metabolism is analyzed through a full-view bright-field imaging method, and the detection result of the drug for treating bladder cancer lymph node metastasis is obtained. Compared with a traditional ENR organ-like culture medium, the method has the advantages that the effect is better, and the cost is lower; the kit is used for screening medicines for bladder cancer lymph node metastasis, is simple and efficient, and can be used for accurately detecting medicines for efficiently killing bladder cancer lymph node metastasis cells.

The invention relates to the technical field of biotechnology or biomedical engineering, in particular to hydrogel, bio-ink, organoids constructed by using the hydrogel, a drug screening model and a construction method. The hydrogel is prepared from 3 to 4 weight percent of gelatin, 0.5 to 2 weight percent of sodium alginate, 20 to 40 weight percent of matrigel and a cell culture medium. The hydrogel is helpful for improving the bionic degree of a tumor microenvironment, can be used for preparing bio-ink containing target cells such as tumor cells, particularly can be used for preparing bio-ink of primary cells (such as primary renal cell carcinoma cells), and an organ-like model constructed by using the bio-ink under a proper 3D biological printing condition is uniform in growth and high in biocompatibility. The cell growth speed is high, the consistency with parent tissues is high, the in-vitro amplification time of the patient-derived tumor cells can be shortened, the influence of subculture factors on the cell affinity can be reduced, the drug development progress can be accelerated, and the production cost can be reduced.

The present invention discloses a method for culturing vitreoretinal organoids in vitro, comprising the following steps: (1) amplifying and culturing pluripotent stem cells to a cell confluence of 50% to 90%; (2) on day 0, digesting the stem cells with a digestion solution; adding a neurosphere culture medium for culturing; adding matrigel for culturing, and then adding a neurosphere culture medium for culturing to obtain a cell suspension; (3) adding a ROCK pathway inhibitor to the cell suspension, and culturing in the neurosphere culture medium for 3 to 5 days to obtain ectoderm neurospheres; (4) replacing the neurosphere culture medium with a vitreous culture medium I, adding TGFB1 protein, and placing the cell suspension on a shaker for 25 to 28 days; (5) replacing the vitreous culture medium I with a vitreous culture medium II, and culturing the cell suspension for 60 to 300 days to obtain vitreoretinal organoids. The vitreous organoids of the retina cultured in vitro according to the present invention can simulate the vitreous in vivo.

The invention relates to a lung cancer organoid and peripheral blood source immune cell co-culture model and a construction method thereof. Specifically, the invention provides a construction method of a macrophage and tumor organoid co-culture model for evaluating anti-tumor activity, monocytes of autologous or allogeneic peripheral blood of a patient are induced and differentiated into high-purity macrophages in vitro, and the high-purity macrophages and lung cancer tumor organoid are subjected to three-dimensional co-culture in matrigel; the model aims at highly reducing a core interaction network of tumor cells and macrophages in TME; the dynamic change of the polarization state of the macrophage is simulated and observed; the problem that an organ-like model is incomplete due to immune component deficiency or spatial positioning distortion of an existing model is effectively solved, an experimental platform closer to the physiological state is provided for tumor immune microenvironment research, and therefore development of the tumor immune treatment field is promoted.

The invention discloses a carbon dot, hydrogel containing the carbon dot as well as a preparation method and application of the hydrogel, and relates to the technical field of carbon materials. The carbon dot is prepared from the following raw materials: a hydroxyl-containing flavonoid compound and epsilon-polylysine; the carbon dots contain aniline bonds formed by hydroxyl groups of the hydroxyl-containing flavonoid compounds and amino groups of the epsilon-polylysine. The carbon dot shows higher tumor cell killing property and lower normal cell toxicity than the raw material hydroxyl-containing flavonoid compound at the cellular level, can obviously induce immunogenic death of cancer cells and does not induce up-regulation of extracellular matrix collagen, and also has a better effect of regulating a tumor immune microenvironment.

The invention discloses a construction method and application of an animal model for evaluating cardiotoxicity of antitumor drugs, sodium carboxymethyl cellulose (CMC-Na) with extremely low cost is adopted to replace matrigel, a tumor-bearing mouse model with high stability can be constructed at low cost, and experimental verification shows that 4T1 cells are resuspended by using a 0.7% CMC-Na solution, and the tumor formation rate of subcutaneous inoculation reaches gt; 95%. The tumor microenvironment-mediated cardiac injury critical period is accurately captured, and the constructed tumor-bearing mouse model is the optimal tumor-bearing mouse model for evaluating the cardiotoxicity of an antitumor drug (adriamycin) in the tumor microenvironment in the third week after the mouse has a 50mm < 3 > tumor.

The invention discloses a preparation method of a mouse skin organoid, which comprises the following steps: separating dermis and epidermis of an E18.5 mouse to obtain dermis cells, mixing the dermis cells with back skin cells of an E14.5 embryo mouse, mixing by using Matrigel to form cell spheres, and gradually forming the skin organoid in in-vitro culture; the organoid is mainly applied to batch production of organoid containing hair follicles. The invention relates to the technical field of stem cells and regenerative medicine. According to the preparation method of the mouse skin organoid, a cell source is improved in the experiment process, the mouse skin organoid containing hair follicles can still be formed after cells in the early development stage are added, and related developmental biology research is carried out. And the source of another dermal cell is not limited to late pregnancy, and also can be a newborn mouse, so that the source of seed cells is expanded, and the cost is reduced.

The invention relates to a co-culture microsphere model with a core-shell structure and a preparation method of the co-culture microsphere model. The co-culture microsphere model comprises a core-shell microsphere, the core-shell microsphere is composed of an inner core and an outer shell, the inner core is an organoid loaded by matrigel, the outer shell is formed by crosslinking a photo-crosslinking hydrogel material, co-culture cells are doped in the photo-crosslinking hydrogel material, and the inner core is wrapped by the outer shell. According to the co-culture microsphere model, double-layer structure microspheres of co-culture cell wrapped organoids are formed, immune infiltration and in-vivo characteristics can be simulated, and a similar in-vivo three-dimensional tumor immune microenvironment is provided. And the types of cells loaded on the inner layer and the outer layer can be flexibly replaced to form a microsphere model for co-culture of single or multiple cells and organoids. The co-culture microsphere model is more stable in structure, the size of the diameter of the core-shell microsphere can be optimized by changing 3D printing parameters, the operability of subsequent research is high, and the co-culture microsphere model is suitable for different research application scenes.

The invention discloses a composition and a method for constructing ovarian organoid based on a clinical tissue sample. The composition comprises a sample preservation solution, a sample digestive juice and a sample culture medium, the method comprises the following operation steps: transporting a preservation solution A to a laboratory, and pretreating to obtain tissue fragment precipitates; carrying out resuspension incubation on the tissue fragment precipitate by using digestive juice B for digestion; collecting tissue blocks which are not completely digested and filtered, and performing secondary digestion to obtain single cells; combining the cells obtained in two times to obtain the required single cell; adding matrigel, then adding the mixture into a pore plate, and putting the pore plate into an incubator until the matrigel is solidified; and taking out the pore plate, adding a culture medium L2, and culturing to obtain the ovarian organ constructed based on the clinical sample. According to the method, the ovarian organoid is efficiently constructed from the ovarian tissue sample; the provided culture solution formula makes up the dilemma that normal ovarian tissue organoids are difficult to form and normal tissue-derived organoids are difficult to passage at present, and reduces the burden and experiment cost brought by sampling in the disease research process.

The invention belongs to the technical field of bioengineering, and relates to heparan sulfate defect type matrigel as well as a preparation method and application thereof. The preparation method comprises the following steps: providing natural matrigel; providing immobilized heparinase, wherein the immobilized heparinase can specifically catalyze and degrade heparan sulfate in the natural matrigel; carrying out contact reaction on the natural matrigel and the immobilized heparinase; and after the reaction is finished, separating the immobilized heparinase. According to the preparation method, the characteristics that the immobilized enzyme is convenient to separate and recycle are utilized, the problems that in a traditional solution enzyme method, enzyme residues exist, the reaction is difficult to accurately control, and subsequent purification steps are complex are solved, and a standardized heparan sulfate defective matrigel product which is controllable in component, free of residual enzyme and high in stability is provided. The method has wide application value in the fields of tumor mechanism research, anti-tumor drug screening, tissue engineering and the like.

The invention relates to the technical field of biology, and discloses a construction method of a gastric cancer organoid culture system, which comprises the following steps: preparation of a conditioned medium: culturing L-WRN cells and collecting the conditioned medium; preparing a gastric cancer organoid culture medium, and mixing the conditioned culture medium, the basic culture medium, a plurality of growth factors and additives; and extraction and culture of organoid: separating cells from gastric cancer tissues, mixing the cells with matrigel, and culturing by using the prepared culture medium. According to the method, the repeatability and the stability of the organoid model are improved, the response consistency of the organoid in drug screening is improved, and the reliability of the model in disease mechanism research and personalized medical application is guaranteed.

This invention relates to a method for constructing a mouse orthotopic model of highly aggregated glioblastoma, belonging to the fields of biomedicine and experimental animal model technology. Glioblastoma cells are resuspended in a composite carrier of a specific ratio of matrix gel and serum-free culture medium, and a low-temperature injection technique is used. Utilizing the thermosensitivity of matrix gel in the mouse intracranial environment (i.e., its phase transition from liquid to gel at a certain temperature), precise colonization and physical locking of tumor cells at target coordinate points are achieved. This method improves upon existing glioblastoma mouse orthotopic model construction techniques that suffer from tumor cell diffusion and loss, low tumor formation rate, irregular tumor morphology, large intragroup variability, and needle reflux due to low viscosity of the inoculation carrier. It provides a good disease model for subsequent screening or evaluation of anti-glioblastoma drugs.

The present invention relates to a preparation method of a composition for culturing lung organoids, a composition thereby, and an organoid culture method using the same. The present invention is capable of creating an environment that is more similar to an actual tissue than a conventional MATRIGEL®-based culture system, and in particular, exhibits an effect of facilitating tissue differentiation in lung organoid culture and effectively developing into a form that is similar to an actual tissue.

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 discloses a construction method and application of a human lung 3D cell microsphere and a human pulmonary fibrosis 3D organoid. The construction of the human lung 3D cell microspheres comprises the following steps: respectively digesting human primary fibrosis cells and human II-type alveolar epithelial cells which are subjected to adherent culture into single cells; mixing digested human primary fibrotic cells, human II-type alveolar epithelial cells and THP-1 macrophages subjected to suspension culture according to a cell number ratio of 1: 1: 1, so as to obtain a cell precipitate; and preparing the cell precipitate into cell matrigel microspheres by using a T-type microfluidic technology, and culturing to obtain the human lung 3D cell microspheres. And further inducing the human lung 3D cell microspheres by using a human lung organoid culture medium containing 50ng / mL of TGF-beta for 24-48 hours to obtain the human pulmonary fibrosis 3D organoid. According to the invention, 3D cell microspheres with human normal lung cell characteristics and protein expression can be formed within 3 days, and pulmonary fibrosis 3D organs can be obtained after short-time induction.

The invention discloses matrigel for brain organ differentiation as well as a preparation method and application of the matrigel. The matrigel provided by the invention is prepared from the following components in percentage by volume: 20 to 40 percent of GFR (Growth Factor Receptor), 50 to 70 percent of I-type collagen, 0.5 to 2 percent of beta-mercaptoethanol and 8 to 9.5 percent of Neurobase + B27. The matrigel provided by the invention can improve the differentiation efficiency of brain organs, improve the stability of an experimental system and reduce the cost.

The invention relates to the technical field of organs, and discloses a culture medium and a method for constructing and culturing vascular organs and application of the culture medium and the method. By utilizing the culture medium set provided by the invention, the vascular organoid with complete functions and structures can be constructed through iPSC induction, and matrigel does not need to be adopted in the construction and culture process, so that the construction operation of the organoid is greatly simplified, and the cost is saved. The blood vessel organoid constructed by the invention has a basic structure of a blood vessel, has corresponding response to medicine components which are known in the field and have influence on the blood vessel, and can be used as a blood vessel in-vitro model.

The application discloses a kind of middle ear organoids culture medium and culture method, belong to organoids technical field.The middle ear organoids culture medium includes: basic culture medium, specific additive factor, CHIR99021 and TTNPB;Specific additive factor includes the following end concentration components: SB203580, 1-20 mmol / L;SB431542, 50-1000 nmol / L;Y-27632, 1-20 mmol / L;FGF-10, 10-30 μg / L;RS-246204, 1-20 nmol / mL and the like.Culture method includes: middle ear tissue is handled to obtain middle ear cell, after middle ear cell is resuspended, with matrigel is mixed, then inoculation, after it coagulates, the above-mentioned middle ear organoids culture medium is added and is cultivated, obtains middle ear organoids.The culture medium of the application can simultaneously improve the quantity and survival rate of middle ear organoids, conducive to industrialized production.

The invention relates to the technical field of biology, in particular to a method for efficiently obtaining human definitive endoderm cells in vitro, which comprises the following steps: S1, inoculating human pluripotent stem cells into matrigel, and performing maintenance culture for 16-24 hours by using a stem cell maintenance culture medium; s2, from Day-2 to Day-1, absorbing and abandoning the stem cell maintenance culture medium, adding a pretreatment culture medium for culture, and changing the pretreatment culture medium once every day; the pretreatment culture medium is a stem cell maintenance culture medium added with dimethyl sulfoxide; s3, carrying out Day0 to Day2, absorbing and abandoning the pretreatment culture medium, adding a definitive endoderm differential culture medium, carrying out induced culture, and changing the definitive endoderm differential culture medium once a day; the definitive endoderm differential culture medium is an RPMI1640 culture medium added with fetal calf serum and activin A; s4, Day3, obtaining the required human definitive endoderm cells, and solving the problems that the existing definitive endoderm differentiation efficiency and cell population purity are limited and are influenced by batch-to-batch variation.