Introduction to cell co-culture experiment

1. Basic principles

In vitro cell co-culture is the co-culture of two kinds of cells (which can come from the same tissue or from different tissues), so that the morphology and function of one of the cells can be stably expressed and maintained for a long time. . This technology can simulate the microenvironment generated in the body, facilitate the better observation of the interaction between cells and cells, cells and culture environment, and explore the mechanism of action and possible targets of the drug, filling the monolayer cell culture and the whole animal. The gap in the experiment.
At present, there are mainly three aspects of research: a. Cell-cell contact and action, causing direct contact with the cytoplasmic membrane and producing extracellular matrix ECM, which contains interstitial collagen, fibronectin, proteoglycan and layer. Components such as fibronectin mimic the microenvironment similar to the body, maintain the stereo configuration of cells and promote cell function. b. Interaction between cells and cells, observe changes in function, traits and growth behavior after cell co-culture, such as the effect of secretion of one cell on the expression of another cell gene, between cells and cells Conversation, etc. c. Study the effects of drugs on cell morphology and function and related mechanisms, and provide important technical support for the development of new drugs.
Cell co-culture is divided into direct contact co-culture and non-direct contact co-culture. Direct contact co-culture refers to co-cultivation of two cells in the same culture dish in a certain ratio under suitable conditions. The main disadvantage of using two cell or tissue contacts, through paracrine, autocrine secreting cytokines or direct contact, is that the separation of the two cells is difficult, and it is difficult to observe and follow-up; non-direct contact co-culture refers to One of the effects of the co-culture system on the other is through paracrine cytokine interactions, but the two do not. Since the two cells are easily separated and are easy to observe and do not affect subsequent detection, the following four methods are included: (1) Coculture with another cell using a culture supernatant of a cell (containing different growth factors). (2) Cells B cultured on a slide (pretreated with type 1 collagen gel) were placed in a Petri dish of Cell A at a certain ratio to be co-cultured. (3) Millicell insert cell culture dish (Transwell chamber) is a kind of transparent cup-shaped device. The bottom layer of the cup is placed with a transparent membrane. The commonly used polycarbonate membrane has a pore size of 0.1 to 12.0. Um. Put the Transwell chamber into the culture plate, the cell A is planted in the upper chamber, and the cells in the lower layer of the cell B can affect the cells in the upper chamber, so that the substance secreted or metabolized by the cell B can be studied for the cell A. influences. (4) Three dimensional cell culture (TDCC): TDCC is a method in which a vector having different three-dimensional structures is co-cultured with various kinds of cells in vitro to enable cells to differentiate to produce a certain three-dimensional tissue-specific structure. A three-dimensional cell carrier complex, such as matrigel, which is commonly used nowadays, is a substance extracted from mouse tumor tissue. At room temperature, it forms a gel-like structure similar to the extracellular basement membrane in vivo, and the endothelium is formed. Within the cell culture Matrigel gel, endothelial cells are characterized by neovascularization. The limitation of the Matrigel gel method is that it does not mimic the microenvironment of tumor and vascular endothelium in vivo. Cell separation and extraction are difficult, and the dispersion of cytokines is affected to some extent.

Ibidi co-culture product introduction ----μ-Slide 2- well co-culture
2. Scope of application: (1) Co-culture of different cell lines or primary cells (2) Interaction of epithelial-mesenchymal cells (3) Paracrine interaction of different types of cells in vitro (4) Cell or cell spheroid culture in Matrigel
3. Features:
(1) IBIDI has 2 well co-cultured slides, the cells do not touch each other, but share the same liquid environment, the cells can be observed in real time, unlike transwell co-culture, the cells need to be taken out to observe;
(2) IBIDI co-culture slides are added to the wells for liquid exchange, culture and observation, etc., which is easier to operate than transwell;
(3) This product can adjust the ratio of donor cells to recipient cells according to experimental needs;
(4) Introduction to product usage: Seed cells are planted in shallow wells. After the cells are attached to the wall, all the shallow wells are immersed in the medium, and the co-culture experiment begins;
- (If the experiment is to do a small number of cells co-culture, or multi-cell co-culture of small cells) IBIDI these 2-hole plug-ins, as well as a 4-hole plug-in can be done, planting fewer cells in the hole of the plug-in;

4. Introduction to the experimental process
1. Seed 40-60 ul of the receptor cell suspension into the middle compartment of the slide, controlled at 5-10 x 104 cells/ml depending on your cell type density;
2. Seed 40-60 ul of feeder layer cell suspension into the other 8 chambers of the slide;
3. After the cells are attached, remove the chamber culture medium and elute the unattached cells, and add 400-600 ul of medium to the large well. The two cells share the same culture system.


Longitudinal section: