one of the techniques that I use in the lab the most often in culturing cells. this is also the technique that I encountered most often when analyzing studying biochemistry. it is also one of the hardest one to talk about because there are so many different variations on it. One of the biggest differences between different cell lines is the media used. Depending on the different cell lines used, there are different media used and different things that are added to cells. While immortalized cells need very few things, only FBS that provides nutrients and antibiotics to prevent contamination, primary normal cell lines can be incredibly fussy, with things like growth hormones and other things that are needed to keep them alive. Making the media itself can be a bit of a challenge because there is a need for many different things at specific concentrations. to cultivate a cell there is a need for a flask or a plate. while some cells are able to grow in a suspension, others must be grown in a container with a surface that the cells can adhere to. the cells are then left in the incubating, which is typically at 37 degrees to grow, however, if the cells are allowed to grow for too long, they will become too confluent, and as a result, the properties of the cell will change. This may or may not be desirable depending on the experiment that the cells are needed for. If the cells cannot be confluent, the cells would need to be split. to do this, the media is removed and the cell surface is washed with PBS and trypsinized so that the cells can detach (if the cell is an adherent cell). Then trypsin is neutralized using media and put into the centrifuge tube to be spun. after it is spun, the media is removed, and new media is added. then the cells are resuspended in the tube and a portion of it is put into a new flask to allow for growth along with the new media that is added.
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