Standardized culture medium and culture method for three-dimensional culture of lung and lung cancer tissue organoids
A three-dimensional culture and culture medium technology, applied in biochemical equipment and methods, culture process, tissue culture, etc., can solve the problems of lack of standardized culture medium, achieve the maintenance of tissue cell specificity, appropriate nutritional components, and meet the needs of scientific research Effect
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Embodiment 1
[0076] A standardized culture medium for three-dimensional culture of lung and lung cancer tissue organoids, including at least one or more of Reagent A and Reagent B;
[0077] Reagent A contains one or more of R-spondin1, Noggin, N-acetylcysteine (N-acetylcysteine), Nicotinamide (Nicotinamide), Y-27632, A8301, SB202190, FGF-7 and FGF-10 ;
[0078] Reagent B comprises reagent B1, reagent B2 and reagent B3;
[0079] Reagent B1 contains Bovine Pitituary Extract (BPE), Y-27632, DAPT, Hydrocortisone, Insulin, Gentamicin, Amphotericin ), Retinoic Acid, Transferrin, Triiodothyronine, Epinephrine and human epidermal growth factor (hEGF);
[0080] Reagent B2 contains heparin (Heparin);
[0081] Reagent B3 contains one or more of DMEM / F12, L-ascorbic acid-2-phosphate magnesium and sodium selenium.
[0082] Preferably, in reagent A, based on the final volume of the standardized medium, the concentration of R-spondin1 is 10%, the concentration of Noggin is 10%, and the concentratio...
Embodiment 2
[0139] This example provides a standardized medium for three-dimensional culture of lung and lung cancer tissue organoids. The standardized medium is used to establish human lung organoids. The formula of the standardized medium is as follows:
[0140] Basal medium:
[0141] Use Advanced F12 / DMEM as solvent and contain the following ingredients:
[0142] N-(2-hydroxyethyl)piperazine-N’-2-ethanesulfonic acid 10~1000mM (specifically, 100mM)
[0143] L-alanyl-L-glutamine 20~2000mM (specifically 200mM)
[0144] Penicillin 10000U / ml
[0145] Streptomycin 10000ug / ml;
[0146] Reagent A:
[0147] Calculated based on the final volume of standardized medium containing the following components:
[0148] R-spondin1 10%
[0149] Noggin 10%
[0150] N-acetylcysteine 0.125~12.5mM (specifically, 1.25mM)
[0151] Niacinamide 1~100mM (for example, 10mM)
[0152] Y-2763 2.5~250uM (specifically 25uM)
[0153] A8301 50~5000nM (for example, 500nM)
[0154] SB202190 0.1~10uM (for examp...
Embodiment 3
[0186] This example provides a standardized medium for three-dimensional culture of lung and lung cancer tissue organoids. The standardized medium is used to expand human lung organoids. The formula of the standardized medium is as follows:
[0187] Basal medium:
[0188] Use Advanced F12 / DMEM as solvent and contain the following ingredients:
[0189] N-(2-hydroxyethyl)piperazine-N’-2-ethanesulfonic acid 10~1000mM (specifically, 100mM)
[0190] L-alanyl-L-glutamine 20~2000mM (specifically 200mM)
[0191] Penicillin 10000U / ml
[0192] Streptomycin 10000ug / ml;
[0193] Reagent A:
[0194] Calculated based on the final volume of standardized medium containing the following components:
[0195] R-spondin1 10%
[0196] Noggin 10%
[0197] N-acetylcysteine 0.125~12.5mM (specifically, 1.25mM)
[0198] Niacinamide 1~100mM (for example, 10mM)
[0199] Y-2763 2.5~250uM (specifically 25uM)
[0200] A8301 50~5000nM (for example, 500nM)
[0201] SB202190 0.1~10uM (for example,...
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