Cell splitting is the process in which a cell line is moved to a different flask in order to keep the cell line alive. Cell splitting must be done before the cell population gets too large. If the cell population gets too big, the cells will overcrowd the flask they are in and die as a consequence. In the bioimaging lab, a course I am currently taking. In this class, my lab partner and I are working with a line of LLC-Pk1 epithelial cells., and currently have three flasks of cells for our project. When the time comes for splitting, we follow a specific protocol. First, we prepare the new flask that the cells are going into with medium and correct labeling. We must then remove the medium that is already in the flasks. After that we wash the cells with about 1 mL of warm PBS (phosphate-buffered saline) and after rocking the flask and making sure the PBS has washed thoroughly, the PBS is removed and cells that have died have been removed from the flask. The next step is to add about 0.5 mL of trypsin, and then incubate the flask for about 3 minutes at 37 degrees Celsius. This process removes cells from the bottom of the flask so that they can now be transferred to the next flask. The trypsin reaction is stopped b adding about 1 mL of cold medium to the flask and triturating thoroughly Finally, we can now put 5 or so drops from the old flask into the new flask, where the cells now have more room to grow until they need to be split again.
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