Methods for culturing cells and uses thereof
AssigneeAcrocyte Therapeutics Inc.
InventorsYing-Chih Chang, Jia-Yang Chen, Yen-Jang Huang
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Abstract
Provided is a method for culturing a sample of cells, which includes circulating epithelial cells, peripheral blood mononuclear cells (PBMC), or body fluid-derived cell samples (e.g. blood, ascites, pleural effusions, cerebrospinal fluids, perfusion fluids, sputum, and urine), or liquid biopsy-derived cell samples in presence of Bone Morphogentic Protein (BMP) inhibitor, a mitogenic growth factor, and a Wnt agonist, when culturing the circulating epithelial cells and relevant blood cells. Also provided is use of cell medium having a BMP inhibitor, a mitogenic growth factor, and a Wnt agonist in obtaining a cell culture of circulating epithelial cells, PBMC and body fluid-derived cells.
Claims17
An in vitro method for obtaining cultured circulating epithelial cells, the method comprising:
- providing a sample of cells including circulating epithelial cells, peripheral blood mononuclear cells (PBMCs), or body fluid-derived cells, and
- culturing the sample of cells in a cell medium for a length of time sufficient to observe formation of one or more cell spheroids having a diameter greater than 20 μm, wherein the cell medium comprises:
- a Bone Morphogenetic Protein (BMP) Inhibitor, and
- a mitogenic growth factor, and
- a Wnt agonist, and
- obtaining, from the cultured cell medium, the one or more cell spheroids containing the cultured circulating epithelial cells.
An in vitro method for obtaining cultured circulating epithelial cells, the method comprising:
- providing a sample of cells including circulating epithelial cells, peripheral blood mononuclear cells (PBMCs), or body fluid-derived cells, and
- culturing the sample of cells in a cell medium for a length of time sufficient to be able to observe formation of one or more cell spheroids having a diameter greater than 20 μm, wherein the cell medium comprises:
- a Bone Morphogenetic Protein (BMP) Inhibitor, and
- a mitogenic growth factor EGF between 0.1 and 500 ng/ml, and
- any one or a combination of Wnt agonist includes Wnt3, Wnt5, R-spondin 1, R-spondin 2, R-spondin 3 and R-spondin 4, Norrin, GSK inhibitor between 0.1 and 3000 ng/ml, and
- a B27 supplement and/or N2 supplement,
- obtaining, from the cultured cell medium, the one or more cell spheroids containing the cultured circulating epithelial cells.
An in vitro method for obtaining cultured circulating epithelial cells, the method comprising:
- providing a sample of cells including circulating epithelial cells, peripheral blood mononuclear cells (PBMCs), or body fluid-derived cells, and
- culturing the sample of cells in a cell medium for a length of time sufficient to be able to observe formation of one or more cell spheroids having a diameter greater than 20 μm, wherein the cell medium comprises:
- a Bone Morphogenetic Protein (BMP) Inhibitor between 0.1 and 500 ng/ml, and
- a mitogenic growth factor including EGF between 0.1 and 500 ng/mL, and
- any one or a combination of Wnt agonist includes Wnt3, Wnt5, RSPO1 to RSPO4 between 1 and 3000 ng/ml, and
- a B27 supplement and/or an N2 supplement
- obtaining, from the cultured cell medium, the one or more cell spheroids containing the cultured circulating epithelial cells.
Dependencies are inferred from the claim text.