10 results about "Dispase" patented technology

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 cell biology, and particularly discloses an isolated culture method of primary meibomian gland epithelial cells, the obtained primary meibomian gland epithelial cells and application of the primary meibomian gland epithelial cells. The method comprises the following steps: efficiently and mildly digesting meibomian gland tissues by adopting a compound digestive juice consisting of I-type collagenase, dispersing enzyme and trypsin in a specific ratio; stopping digestion by using a serum-containing neutralizing solution and collecting cells; the method comprises the following steps: promoting cell attachment by using a serum-containing initial culture medium, and carrying out selective amplification by using a serum-free proliferation culture medium to finally obtain high-purity and high-activity primary meibomian gland epithelial cells. The method solves the problems of long digestion time, low cell yield, poor purity and the like in the traditional method. The obtained cell can be used for constructing a meibomian gland dysfunction model, screening drugs and researching lipid metabolism, and an important technical platform and a cell model are provided for research and treatment of related eye diseases.

The invention relates to a method for culturing adult hair follicle protuberance cells, which belongs to the technical field of biology and comprises the following steps: S1, cleaning hair follicle tissues to obtain hair follicle tissues containing protuberance cells; s2, the hair follicle tissue containing the protuberant cells is soaked in an Advanced DMEM / F12 culture medium mixed solution containing collagenase, dispersing enzyme, EGF, bFGF and 1 * green-streptomycin, and digestion is carried out; s3, chopping hair follicle tissues containing the protuberant cells, centrifuging after stopping digestion, adding a cell culture dissociation agent into precipitated tissues, digesting twice, collecting supernatant after stopping digestion, centrifuging, collecting precipitates, re-suspending, and filtering; and S4, inoculating the cells into a cell six-well plate for 3D suspension culture, and changing the solution and blowing and beating the cells every other day. According to the method, a two-step method is adopted to digest the tissue part of the hair follicle convex cells, so that a larger number of hair follicle convex cells are obtained, and the characteristics and dryness of the hair follicle convex cells are improved.

The present invention relates to a method for non-destructively peeling off a cell sheet in culture from a cell culture surface of a culture container, belongs to the field of biotechnology, and comprises the following steps: S1, preparing a digestion solution: mixing DMEM / F12 culture medium, dispase, collagenase, and ROCK inhibitor to obtain a digestion solution; S2, cell sheet digestion: adding a cell sheet to be peeled off to the digestion solution obtained in step S1, and incubating; S3, peeling off a cell sheet: affixing the cell sheet to be peeled off to a silicone gauze, then simultaneously clamping the cell sheet-silicone gauze and pulling it away from one end of a cell culture dish until it is separated from the cell culture surface of the cell culture dish, obtaining a cell sheet-silicone gauze, that is, completing the rapid and non-destructive separation of the cell sheet in culture from the cell culture surface of the culture dish. Compared with the prior art, the method of the present invention can make the cell sheet completely detached, and the bottom of the culture dish has less adhesion; the overall operation is convenient, the time is short, and it is suitable for the preparation and peeling off of a large number of cell sheets.

The invention discloses a kit and a method for preparing alligator sinensis tissue single cells, and belongs to the technical field of single cell sequencing. The kit comprises Collagenase I, DNase I, Dispase and a DMEM (Dulbecco Modified Eagle Medium) culture medium. The kit and the method provided by the invention fill the blank of the Yangtze alligator tissue single cell preparation technology. The kit is simple in component and easy to prepare, the method has very high repeatability and success rate, a large number of high-motility and low-impurity single cells can be obtained, and the kit can be applied to single cell sequencing and more application fields.

The application belongs to the technical field of biological medicine, and particularly relates to a thymus tissue dissociation reagent, a kit and application thereof in obtaining thymus epithelial cells. The thymus tissue dissociation reagent at least comprises collagenase D, dispersinase II, deoxyribonuclease I, dissociation enzyme and a dissolving reagent; and collagenase II and / or collagenase IV and / or trypsin. The cell survival rate of the thymus tissue dissociation reagent after dissociating the thymus tissue is maintained at a high level of 95.7% to 99.6%, which is significantly higher than that of the prior art; and after sorting, the number of thymus epithelial cells enriched and obtained from each gram of thymus tissue is 1.38e 5 ~1.06e 6 , and the success rate of enriching and obtaining thymus epithelial cells is as high as 95%, which is significantly higher than that of the prior art; and the proportion of epithelial stem cells in the thymus epithelial cells is 65.7%, which is significantly higher than that of the prior art.

The invention discloses a combined reagent for separation of keloid macrophages and application. The combined reagent for separation comprises a first buffer solution and a second buffer solution, the first buffer solution comprises enzyme A, and the enzyme A is dispersing enzyme; the second buffer solution comprises enzyme B, and the enzyme B comprises collagenase I, collagenase II, collagenase IV, collagenase V, hyaluronidase and deoxynuclease; the application refers to the application of the combined reagent for separation in preparation of keloid macrophage separation products. According to the invention, the separation efficiency and the cell survival rate of the macrophages are obviously improved.

The invention particularly relates to a human pneumosarcoma-like carcinoma organ as well as a construction method and application thereof, and belongs to the technical field of biological medicines. The organoid is selected from TJLCI-PSC-O-1 with the preservation number of CCTCC (China Center For Type Culture Collection) NO: C2025218 or TJLCI-PSC-O-2 with the preservation number of CCTCC NO: C2025334, or an amplified culture of an epithelial marker CK and a mesenchymal marker VIMENTIN which are stably co-expressed by the organoid. According to the construction method, a collagenase / dispersing enzyme and DNase I combined stepped enzymolysis system and a three-dimensional culture system are adopted, and high yield and long-term stable amplification of cells are realized. The model highly simulates the pulmonary sarcomatoid carcinoma pathological structure and epithelial-mesenchymal transition phenotype for the first time, can be used for in-vitro drug sensitivity test, in-vivo tumor formation model construction and drug screening, and provides a key platform for basic research and precise treatment of diseases.

This invention discloses a kit and method for preparing single cells from reptiles, belonging to the field of single-cell sequencing technology. The kit includes Collagenase I, DNase I, Dispase, and DMEM culture medium. The kit and method of this invention fill the gap in single-cell preparation technology from reptile tissues, especially the Chinese alligator. The kit has simple components and is easy to prepare; the method has high reproducibility and success rate, and can obtain large quantities of single cells with high viability and low impurities, which can be used for single-cell sequencing and other applications.

The present invention relates to a method for culturing adult hair follicle bulge cells, belonging to the field of biotechnology, and comprising the following steps: S1. Cleaning and processing the hair follicle tissue to obtain the hair follicle tissue containing bulge cells; S2. Immersing the hair follicle tissue containing bulge cells in a mixed solution of Advanced DMEM / F12 medium containing collagenase, dispase, EGF, bFGF, and 1× penicillin-streptomycin for digestion; S3. Chopping the hair follicle tissue containing bulge cells, centrifuging after terminating digestion, adding a cell culture dissociating agent to the precipitated tissue for digestion twice, centrifuging the supernatant after terminating digestion, collecting the precipitate, resuspending and filtering; S4. Inoculating the cells into a six-well cell plate for 3D suspension culture, changing the medium every other day and pipetting the cells. The present invention uses a two-step method to digest the hair follicle bulge cell tissue site, obtaining a larger amount of hair follicle bulge cells and improving the characteristics and stemness of the hair follicle bulge cells.