Organoid culture system and organoid culture method
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A culture system and organ culture technology, applied in the field of organoid and cell culture, can solve the problems of unfavorable large-scale culture and application of organoids, complicated culture steps, high culture cost, etc., and achieve simplified culture process and culture cost, easy conditions, Cultivate effects with simple steps
Pending Publication Date: 2021-12-07
COHERENT BIOPHARMA (SUZHOU) LTD
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Although this method realizes the 3D culture of kidney tissue organoids, it needs to go through a complicated process of tissue disruption and cell separation, which has the problems of complicated culture steps and high culture costs, which is not conducive to the large-scale culture and application of organoids.
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Embodiment 1
[0113] Example 1: 3-D Culture of Kidney Tissue Organoids
[0114] Step 1, 293T cells were revived, cultured with IMDM (10% (v / v) fetal bovine serum), and passaged.
[0115] Step 2: Select 293T cells in a well-cultured and well-grown culture dish, wash them twice with PBS, and digest with 0.25% trypsin. Stop the reaction.
[0116] Step 3, collect the cell suspension into a centrifuge tube, centrifuge at 1000rpm for 5min, discard the supernatant, wash the cells with PBS, then centrifuge at 1000rpm for 5min, repeat washing twice, and then resuspend with 1ml medium cells, and count the cells using a cell counter.
[0117] Step 4, according to the number of cells, dilute the cells with medium to 100 μl system containing 80 cells to obtain a cell suspension for later use.
[0118] Step 5, select Matrigel ( Growth Factor Reduced (GFR) Basement Membrane Matrix, Phenol Red-free, LDEV-free, Life Sciences; 356231) was used as Matrigel, first take 32 μl Matrigel, spread it in a 96-...
Embodiment 2
[0120] Example 2: 3-D Culture of Kidney Tissue Organoids
[0121]The difference between this example and Example 1 is that the matrigel used in step 5 is RGF-BME (CultrexReduced Growth Factor Basement Membrane Extract, Type 2, Pathclear R&D; 3533-005-02).
Embodiment 3
[0122] Example 3: 3-D Culture of Kidney Tissue Organoids
[0123] The difference between this example and Example 2 is that in step 4, the cells are diluted to a 100 μl system containing 800 cells.
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Abstract
The invention belongs to the technical field of organoid and cell culture, and particularly relates to an organoid culture system, an organoid culture method, an organoid obtained based on the organoid culture system or culture method, an application of the organoid of the organoid culture system or the organoid culture method, and an application of the organoid. The organoid culture system or culture method provided by the invention realizes organoid culture by using cells derived from a cell line, can effectively simplify the culture process of the organoid, reduces the culture cost, is suitable for culture and application of organoid models of various tissues, and provides an important research and application platform for the fields of disease model construction, drug screening, drug toxicity testing, individualized treatment and the like.
Description
technical field [0001] The invention belongs to the technical field of organoids and cell culture. Specifically, the invention relates to an organoid culture system, an organoid culture method, organoids obtained based on the organoid culture system or culture method and applications thereof. Background technique [0002] Two-dimensional (two-dimensions, 2-D) cell culture is a commonly used method for in vitro cell culture. The 2-D culture process is simple and efficient, and it is widely used in biomedical research. However, there is a huge gap between the growth environment of 2-D cultured cells and the in vivo environment, which leads to great changes in the biological characteristics and behavior of cells, making it difficult to simulate the three-dimensional microenvironment in vivo. Three-dimensional (three-dimensions, 3-D) cell culture is the co-cultivation of cells with biomaterials with three-dimensional structures. Biomaterials provide growth factors or three-dimen...
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