Neural stem cell preparation for treating Parkinson's disease through nasal delivery
A technology for neural stem cells and Parkinson's disease, applied in the field of neural stem cell preparations and their preparation, can solve the problem of irreversible degeneration of DA neurons, and achieve the effects of good curative effect and safety, repair of nerves and tissues, and stable performance.
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Embodiment 1
[0031] Example 1: Acquisition of Neural Stem Cell Seed Bank Cells
[0032] 1. Induction of primary neural stem cells from embryonic stem cells (ESCs)
[0033] Use a cell scraper to pick ESCs cell clones, blow them gently, and then resuspend them with neural stem cell induction medium, transfer them to an ultra-low adsorption culture dish for suspension culture for about 3 days, collect the formed cell balls, blow them gently, and then use neural stem cell The stem cell induction medium was resuspended, suspended and cultured in an ultra-low adsorption culture dish for 3 days, and the formed cell spheres were collected; the culture dish coated with Laminin was washed twice with DPBS without Ca and Mg one day in advance, and the collected The cell balls were resuspended with the complete medium of neural stem cells, inoculated in the coated culture dish for adherent culture, and the medium was changed every 2-3 days until obvious neural-like colonies were formed, and then the fo...
Embodiment 2
[0044] Example 2: Acquisition of Neural Stem Cell Master Cell Bank Cells
[0045] 1. Coating of culture flasks: add an appropriate amount of laminin Laminin solution to the cell culture flasks, put them in a 37°C incubator for coating for 2 hours, and then wash them once with DPBS without Ca and Mg for later use;
[0046] 2. After digesting and counting the amplified and purified P5 neural stem cells, resuspend them in complete serum-free medium, and adjust the density to 1x10 5 cells / ml, inoculated in coated culture flasks, placed at 35°C 5% CO 2 The adherent culture is carried out in the incubator; the cell culture medium is changed every 48 hours, and the cells can be digested and passaged after continuous culture for about 7 days until the P8 generation. The cells obtained during this period are stored in the main cell bank.
Embodiment 3
[0047] Example 3: Acquisition of Neural Stem Cell Working Cell Bank Cells
[0048] Resuspend P8 passage cells in serum-free complete medium, and adjust the density to 1x10 5 cells / ml, inoculated in coated culture flasks, placed at 35°C 5% CO 2 Adhesive culture is carried out in the incubator. During the culture process, samples are taken to detect cell characteristics and heterologous substances. After passing the test, the cells are frozen and transferred to the working cell bank for later use.
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