Reagent and kit for enriching breast cancer stem cells from breast cancer cell strains
A breast cancer stem cell and breast cancer cell technology, applied in the field of stem cells, can solve the problems of high cost and long cycle, and achieve the effects of improving efficiency, high content and short cell sphere formation cycle
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Embodiment 1
[0024] Example 1: Enrichment of breast cancer stem cells from human breast cancer MCF-7 cell line
[0025] 1. Experimental materials
[0026] Conventional serum-containing medium: 10% fetal bovine serum FBS and double antibody were added to DMEM / F12 culture medium, and 10 ml of penicillin-streptomycin solution (100× double antibody, Beijing Legen Biotechnology Co., Ltd.) was added to 1L medium.
[0027] Conventional serum-free medium: add double antibody (concentration as above), recombinant human epidermal growth factor EGF 20g / L, bovine serum albumin BSA4g / L, insulin 5mg / L, B272% (1L medium plus 20ml 50X B27).
[0028] Improved serum-containing medium: 10% fetal bovine serum FBS, double antibody (same concentration as above), 150nmol / L of galantamine and lycorine were added to DMEM / F12 culture medium, and the molar ratio of the two was 1:1.
[0029] Improved serum-free medium: add double antibody (concentration as above), recombinant human epidermal growth factor EGF 20g / ...
Embodiment 2
[0059] Example 2: Enrichment of breast cancer stem cells from human breast cancer MCF-7 cell line
[0060] 1. Experimental materials
[0061] Improved serum-containing medium: a total of 150nmol / L of galantamine and lycorine, the molar ratio of the two is 2:1, and the others are the same as in Example 1.
[0062]Improved serum-free medium: a total of 150nmol / L of galantamine and lycorine, the molar ratio of the two is 2:1, and the others are the same as in Example 1.
[0063] 2. Experimental method
[0064] With embodiment 1.
[0065] 3. Experimental results
[0066] (1) Formation of MCF-7 microspheres
[0067] Typical MCF-7 suspension cell spheres appeared on the 3rd day, and the cell sphere formation efficiency was (3.1±0.9)%.
[0068] (2) CD44 + CD24 - / low cell ratio
[0069] FCM showed that CD44 in microsphere cells on day 3 of the improved culture method + CD24 - / low The cell ratio was (59.7±5.4)%.
[0070] (3) SP cell ratio
[0071] The proportion of SP cel...
Embodiment 3
[0072] Example 3: Enrichment of breast cancer stem cells from human breast cancer MCF-7 cell line
[0073] 1. Experimental materials
[0074] Improved serum-containing medium: a total of 150nmol / L of galantamine and lycorine, the molar ratio of the two is 1:2, and the others are the same as in Example 1.
[0075] Improved serum-free medium: a total of 150nmol / L of galantamine and lycorine, the molar ratio of the two is 1:2, and the others are the same as in Example 1.
[0076] 2. Experimental method
[0077] With embodiment 1.
[0078] 3. Experimental results
[0079] (1) Formation of MCF-7 microspheres
[0080] Typical MCF-7 suspended cell spheres appeared on the 3rd day, and the cell sphere formation efficiency was (3.3±0.5)%.
[0081] (2) CD44 + CD24 - / low cell ratio
[0082] FCM showed that CD44 in microsphere cells on day 3 of the improved culture method + CD24 - / low The cell ratio was (56.5±5.8)%.
[0083] (3) SP cell ratio
[0084] The proportion of SP cel...
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