Antibiotics are routinely used in cell cultures to prevent possible contamination. However, this use has side effects: many scientific studies show that its use affects cell growth and differentiation. With good laboratory practices, the use of antibiotics is unnecessary.
- Cell culture: a complex microcosm
- Antibiotics deeply influence cell metabolism
- Negative effects of antibiotics on human keratinocytes
- Try to reduce antibiotics use from your cell culture
- Related blog entries
Cell-culture: a complex microcosm
A cell culture is a world of its own. The culture dish contains nutrients within the media that cells need to thrive and prosper. But every now and then, unwanted intruders sneak into the cultures and endanger the fragile balance of the system, spoiling the scientific results. In the end, the cultures are useless and must be discarded. But how can we keep these invaders out? One answer for many scientists is using antibiotics. These compounds are the frontline therapy in the fight against bacterial contamination
These compounds are the frontline therapy in the fight against bacterial contamination. Cell culture is one of the most common and complex technologies in the life sciences and provides an excellent model for investigating physiological and pathological processes, as well as the effects of drugs and toxic compounds at the cellular and subcellular levels. Cell culture is also used for large-scale production of vaccines, therapeutic proteins and cells for regenerative medicine. The media utilized in various laboratory settings represent a critical element for maintaining healthy, proliferating cells. One fundamental requirement of culturing cells in vitro is avoiding microbiological contamination.
This is why standard cell culture protocols often include the prophylactic use of antibiotics, such as penicillin, streptomycin, gentamicin or amphotericin as media supplements to reduce infection rates. Relatively little is known, however, about the effects of these substances on the metabolism of cultured cells, cell proliferation, differentiation or gene expression. Do antibiotics indeed help to solve the contamination issue, or are they creating additional new problems? “The main goal to use antibiotics in cell culture is to kill bacteria or inhibit their proliferation,” explains Dr. Muna Ali, a scientific support specialist at PromoCell. “However, it is easy to forget that they can also harm the cells themselves as they cause many side effects: For instance, antibiotics can attack other specific, non-bacterial structures in the cell.”
Antibiotics deeply influence cell metabolism
From the 1970s to the 90s, numerous reports presented evidence of the antiproliferative effects of beta-lactam antibiotics like penicillin on cultured eukaryotic cells (Neftel et al., 1987). Similar findings were obtained when using aminoglycosides such as streptomycin or gentamicin (Fischer et al., 1975, Cooper at al., 1990, Cooper et al., 1991). Nevertheless, most research groups continued to incorporate antibiotics in their culture media. “One reason why researchers forget about the side effects of antibiotics is that they are not always obvious”.
However, recent advances in regenerative medicine, and the increasing use of cultured cells for therapeutic approaches, have prompted investigators to better evaluate the influence of antibiotics on the biochemistry and differentiation potential of human cultured cells rather than cell lines. Llobet and colleagues described how the mix of penicillin/streptomycin or gentamicin alone can affect the differentiation of human adipose-tissue derived stem cells into adipocytes. Similar effects were observed on embryonic stem cells (Cohen et al., 2006, Varghese et al., 2017), mesenchymal stem cells (Chang et al., 2006), primary cancer cell lines (Relier et al., 2016) and keratinocytes (Nygaard et al., 2015). The use of antibiotics can also significantly alter gene expression and regulation (Ryu et al., 2017) and could modify the results of studies focused on drug response, cell cycle regulation and cell differentiation.
Negative effects of antibiotics on human keratinocytes
Normal human epidermal keratinocytes (NHEK) are widely used in the field of basic research, and conventional monolayer or three-dimensional cultures have been the standard approach for almost 30 years. Beta-lactam antibiotics, as well as aminoglycosides, reduce the proliferation rate of NHEK and hinder the development of a fully differentiated epidermis in 3D skin models (Nygaard et al., 2015).
In a parallel test, NHEK from the same batch were cultured with and without gentamicin and amphotericin under identical conditions in Duesseldorf. The same cell batch was also cultured in the PromoCell laboratories according to their specific guidelines, which recommends avoiding antibiotics in the culture medium.
Try to reduce the antibiotic use from your experiments
In the absence of antimicrobial agents, the best strategy is to work aseptically. In such a way that it is important to make sure to work in a clean, effective and safe way inside the cell culture hood. Try to divide your bell into a clean side (in my case, it used to be the right side) and a dirty side (the left side), in order to minimize any possibility of contamination.
Other tips to take into account, in order to minimize the risk of contamination, is to regularly clean water baths, centrifuges and incubators on a regular basis. It is recommended to wash all these surfaces with WaterShield™ from Minerva Biolabs before cleaning with 70% ethanol.
A fact that is often overlooked is the cleaning of the cell culture hood itself. Sure, we all clean it before we start and after we finish. Is it not like this? But have ayou ever looked under the tray? Do not forget to give the bell a good cleaning every month or so. If you take care of your cells and your home (even in the absence of antibiotics and antifungals), your cells will help youand your results!
Related blog entries
To finish, we leave you these related blog entries that may be of interest:
- Mycoplasma contamination – small organisms cause big troubles
- A home for the cancer stem cells: 3D Tumorspheres
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