Human intestinal cancer primary cells, applications and culture methods
A primary cell and intestinal cancer technology, applied in cell culture active agents, tumor/cancer cells, biochemical equipment and methods, etc., can solve the problems of poor cell state, culture failure and death of primary human intestinal cancer cells
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Embodiment 1
[0075] Example 1: Isolation and culture of human intestinal cancer primary cells
[0076] (1) With the informed consent of the patient or the patient's guardian, collect part of the intestinal cancer tissue removed during the patient's operation;
[0077] (2) Transfer the intestinal cancer tissue collected in step (1) to a cell culture dish, wash with a cleaning solution (containing 0.4 mg / ml penicillin, 0.4 mg / ml kanamycin sulfate, 0.4 μg / ml amphotericin B, 4 μg / ml vancomycin in normal saline) for 8 times to remove non-cancerous tissue impurities such as fat and mucous membrane;
[0078] (3) Transfer the cleaned intestinal cancer tissue in step (2) to a new culture dish, cut the cleaned intestinal cancer tissue into small pieces with scissors and a blade, and chop them into fine pieces;
[0079] (4) Transfer the crushed intestinal cancer tissue to a 50ml centrifuge tube, centrifuge at 2000rpm for 8min, remove the supernatant, add 9ml DF medium for resuspension, and then add ...
Embodiment 2
[0084] Example 2: Purification of Human Intestinal Cancer Primary Cells
[0085] (1) When the confluence of the cells cultured in step (8) of Example 1 reaches 70-90%, the serum-free medium is removed and digested with 1-2 ml EDTA-Trypsin.
[0086] (2) Observe under a microscope until the cells are detached from the culture flask, discard the fibroblasts digested by EDTA-Trypsin first, and then collect the digested cells while digesting, and stop the digestion of the collected cells with DF10 medium until All cells were digested to completion.
[0087] (3) Centrifuge at 1500rpm for 6min, remove the supernatant, resuspend with serum-free medium, and store at 37°C, 5% CO 2Place in the cell incubator for 1.5 hours, after the fibroblasts adhere to the wall, collect the unattached cells, transfer them to a new culture bottle to continue culturing, and repeat the adhesion for 3 times to achieve the effect of completely removing the fibroblasts. The primary human intestinal cancer ...
Embodiment 3
[0089] Embodiment 3: Morphological observation
[0090] By observing under an inverted microscope, it can be found that the human intestinal cancer primary cells cultured after Examples 1 and 2 are in the form of epithelial cells. Cells proliferate rapidly, without contact inhibition, squeeze each other, and appear stacked or mosaic. For specific results, see figure 1 .
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