1970s commercialization of this technology had

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Unformatted text preview: s, commercialization of this technology had further impact on cell culture that continues to this day. Companies, such as Corning, began to develop and sell disposable plastic and glass cell culture products, improved filtration products and materials, liquid and powdered tissue culture media, and laminar flow hoods. The overall result of these and other continuing technological developments has been a widespread increase in the number of laboratories and industries using cell culture today. How Are Cell Cultures Obtained? Primary Culture Fixed and stained human foreskin explants on the surface of a 150 mm culture dish. The explants were cultured for approximately two weeks. Two of the nine explants (bottom left and right corners) failed to grow. The remaining explants show good growth. Each square is approximately 2 cm across. When cells are surgically removed from an organism and placed into a suitable culture environment, they will attach, divide and grow. This is called a Primary Culture. There are two basic methods for doing this. First, for Explant Cultures, small pieces of tissue are attached to a glass or treated plastic culture vessel and bathed in culture medium. After a few days, individual cells will move from the tissue explant out onto the culture vessel surface or substrate where they will begin to divide and grow. The second, more widely used method, speeds up this process by adding digesting (proteolytic) enzymes, such as trypsin or collagenase, to the tissue fragments to dissolve the cement holding the cells together. This creates a suspension of single cells that are then placed into culture vessels containing culture medium and allowed to grow and divide. This method is called Enzymatic Dissociation. Remove tissue Mince or chop Digest with proteolytic enzymes Place in culture Enzymatic Dissociation Subculturing When the cells in the primary culture vessel have grown and filled up all of the available culture substrate, they must be Subcultured to give them room for continued growth. This is usually done by removing them as gently as possible from the substrate with enzymes. These are similar to the enzymes used in obtaining the primary culture and are used to break the protein bonds attaching the cells to the substrate. Some cell lines can be harvested by gently scraping the cells off the bottom of the culture vessel. Once released, the cell suspension can then be subdivided and placed into new culture vessels. Primary culture from the fish Poeciliopsis lucida. Embryos were minced and dissociated with a trypsin solution. These cells were in culture for about 1 week and have formed a confluent monolayer. 2 Once a surplus of cells is available, they can be treated with suitable cryoprotective agents, such as dimethylsulfoxide (DMSO) or glycerol, carefully frozen and then stored at cryogenic temperatures (below -130°C) until they are needed. The theory and techniques for cryopreserving cells are covered in the Corning Technical...
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This note was uploaded on 09/30/2013 for the course PHARM 101 taught by Professor Mishra during the Fall '11 term at Birla Institute of Technology & Science, Pilani - Hyderabad.

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