The practice of keeping cells alive in culture


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Cell culture media and its associated reagents/supplements by ScienCell Research Laboratories. Photo by ScienCell.

The practice of keeping cells alive in culture

Cells can be separated from tissues for ex vivo culture in many ways. Blood cells are easy to separate, but only white blood cells can grow in a lab setting. Cells can be taken out of solid tissues by using enzymes like collagenase, trypsin, or pronase to break down the extracellular matrix, then shaking the tissue to get the cells into suspension. Pieces of tissue can also be put in a growth medium, and the cells that grow out can be used to culture. This technique is referred to as explant culture.

Primary cells are cells that come straight from a person and are grown in a lab. Most primary cell cultures don’t last long, except for those that come from tumors.

An immortalized cell line has the ability to divide indefinitely because of a random mutation or a change made on purpose, like putting the telomerase gene into action. There are a lot of cell lines that have been shown to be good examples of their type.

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Most isolated primary cells go through a process called senescence, which means that after a certain number of population doublings, they stop dividing but remain alive.

Aside from temperature and gas mixture, the cell growth medium is the most often modified part of a culture system. The pH, amount of glucose, growth factors, and other nutrients in a growth medium recipe can be different from one to the next. The growth factors used to complement media are often obtained from the serum of animal blood, such as fetal bovine serum (FBS), bovine calf serum, porcine serum, and equine serum.  One problem with these blood-based ingredients is that viruses or prions could get into the culture, especially when they are used in medical biotechnology.  Alternative strategies include getting animal blood from places with less risk of these diseases, like the United States, Australia, and New Zealand, and using purified nutrient concentrates made from serum instead of whole animal serum for cell culture.

For some types of cells, plating density (the number of cells per volume of culture medium) is very important. For instance, when the plating density is low, granulosa cells make estrogen, but when the plating density is high, they look like theca lutein cells that make progesterone.

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There are two ways to grow cells: suspended or on a surface. Some cells naturally live in suspension, without being attached to a surface, such as cells that exist in the bloodstream. There are also cell lines that have been changed so that they can live in suspension cultures and grow to a higher density than they could in an adhesion environment. Adherent cells need a surface like tissue culture plastic or a microcarrier, which may be coated with extracellular matrix components like collagen and laminin to improve adhesion and send other signals that cells need to grow and change. Most cells that come from solid tissues stick to each other. Organotypic culture is another type of adherent culture. Instead of growing cells in two-dimensional culture dishes, it grows cells in a three-dimensional (3-D) environment. This 3D culture system is more like in-vivo tissue biochemically and physiologically, but it is hard to keep up because of many factors.

Sources:

Voigt N, Pearman CM, Dobrev D, Dibb KM (September 2015). “Methods for isolating atrial cells from large mammals and humans”. Journal of Molecular and Cellular Cardiology. 86: 187–198. doi:10.1016/j.yjmcc.2015.07.006. PMID 26186893

Louch WE, Sheehan KA, Wolska BM (September 2011). “Methods in cardiomyocyte isolation, culture, and gene transfer”. Journal of Molecular and Cellular Cardiology. 51 (3): 288–298. doi:10.1016/j.yjmcc.2011.06.012. PMC 3164875. PMID 21723873

Hemeda, H., Giebel, B., Wagner, W. (16Feb2014) Evaluation of human platelet lysate versus fetal bovine serum for culture of mesenchymal stromal cells Cytotherapy p170-180 issue 2 doi.10.1016

“LipiMAX purified lipoprotein solution from bovine serum”. Selborne Biological Services. 2006. Archived from the original on 2012-07-19. Retrieved 2010-02-02

Portela VM, Zamberlam G, Price CA (April 2010). “Cell plating density alters the ratio of estrogenic to progestagenic enzyme gene expression in cultured granulosa cells”. Fertility and Sterility. 93 (6): 2050–2055. doi:10.1016/j.fertnstert.2009.01.151. PMID 19324349

Jaccard N, Macown RJ, Super A, Griffin LD, Veraitch FS, Szita N (October 2014). “Automated and online characterization of adherent cell culture growth in a microfabricated bioreactor”. Journal of Laboratory Automation. 19 (5): 437–443. doi:10.1177/2211068214529288. PMC 4230958. PMID 24692228

Humpel C (October 2015). “Organotypic brain slice cultures: A review”. Neuroscience. 305: 86–98. doi:10.1016/j.neuroscience.2015.07.086. PMC 4699268. PMID 26254240

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