Hi Yong,

This is a very important issue. I do see both sides of the fence here. In an ideal world, with sparkling white labs that uphold universal standards and best practices, there would be no need for using antibiotics in general cell culture. But this world is far from perfect, and if you have ever seen an undergraduate intern learning cell culture techniques, these laboratory imperfections (the learning curve) is very apparent. Bells and whistles and alarms should sound, PI’s should shriek in horror, but this is a great part of research that deserves as much respect as cell culture; education. I do think it would be best if most general cell culture could be done without the use of antibiotics as antibiotics obviously and undoubtedly contribute to the cellular microenvironment and all results must take into account the use of antibiotics. But unfortunately, many research labs experience breeches in sterility, so the use of antibiotics serves two main purposes. First, it provides intra-laboratory protection to a researcher and allows a researcher to do cell culture with an extra buffer of protection against unknown factors (such as interns). Don’t get me wrong, sterile technique is very important for each and every researcher and if you are using sterile techniques and disinfecting the cell culture hood prior to use, environmental contamination should not be an issue, but there is an increased probability for contamination in educational research environments. The second reason the use of antibiotics in cell culture is beneficial is that it more readily permits the replication of scientific data; it provides inter-laboratory protection. Not all labs are the same and not all labs face the same environmental concerns. Using antibiotics in cell culture offers a layer of protection against the vast environmental variability between labs and more readily allows inter-laboratory replication of cell culture experiments. The dissemination and replication of scientific data is a critical step in the progression of science. Using antibiotics in cell culture aids this data replication process.

Ideally, antibiotics would not be necessary in cell culture, but to protect against intra- and inter-laboratory environmental factors I use a bacteriostatic and bactericidal solution (1% penicillin/ streptomycin/ amphotericin B mixture; Lonza; Walkersville, MD).

Antibiotics are not required for cell growth. It is just a safe guard against bacterial contamination. If you use aseptic conditions during your handling cell cultures, you should not have any problems at all. The exception is the use of antibiotics required for selection of particular cells in a cell culture. I do not use antibiotics at all in cell culture and we have separate room only for cell culture.

Hi Yong,

This is a very important issue. I do see both sides of the fence here. In an ideal world, with sparkling white labs that uphold universal standards and best practices, there would be no need for using antibiotics in general cell culture. But this world is far from perfect, and if you have ever seen an undergraduate intern learning cell culture techniques, these laboratory imperfections (the learning curve) is very apparent. Bells and whistles and alarms should sound, PI’s should shriek in horror, but this is a great part of research that deserves as much respect as cell culture; education. I do think it would be best if most general cell culture could be done without the use of antibiotics as antibiotics obviously and undoubtedly contribute to the cellular microenvironment and all results must take into account the use of antibiotics. But unfortunately, many research labs experience breeches in sterility, so the use of antibiotics serves two main purposes. First, it provides intra-laboratory protection to a researcher and allows a researcher to do cell culture with an extra buffer of protection against unknown factors (such as interns). Don’t get me wrong, sterile technique is very important for each and every researcher and if you are using sterile techniques and disinfecting the cell culture hood prior to use, environmental contamination should not be an issue, but there is an increased probability for contamination in educational research environments. The second reason the use of antibiotics in cell culture is beneficial is that it more readily permits the replication of scientific data; it provides inter-laboratory protection. Not all labs are the same and not all labs face the same environmental concerns. Using antibiotics in cell culture offers a layer of protection against the vast environmental variability between labs and more readily allows inter-laboratory replication of cell culture experiments. The dissemination and replication of scientific data is a critical step in the progression of science. Using antibiotics in cell culture aids this data replication process.

Ideally, antibiotics would not be necessary in cell culture, but to protect against intra- and inter-laboratory environmental factors I use a bacteriostatic and bactericidal solution (1% penicillin/ streptomycin/ amphotericin B mixture; Lonza; Walkersville, MD).

We don't use antibiotics for regular stuff. However for cell sorting experiments we use antibiotics.

I routinely use penicillin and streptomycin for culturing cell lines, or where this is not appropriate I substitute for Gentamicin and Amphotericin.

Some time it has to be done but not routinely.

You do not routinely add antibiotics into culture medium especially when you plan to knock-in or knock-down a protein. When cells are treated with transfection reagent then antibiotics can hurt the cells. Also, you do not add antibiotic when you plan to select a transfected cell clone since some plasmids have put in antibiotic-sensitive gene for clone selection. To some extent antibiotics are toxic to cells like gentamicin can inhibit protein synrthesis. But, you do not want to have cells infected in your incubator either. So, it becomes a judgement call.

Using antibiotics depend on your objective. Less you use better is.

The choice of the antibiotics you will use, will depend on the protection you want to give:

Wide range of action antibiotics

More restricted range of action.

It also depends on which kind of cells your are working primary cells from animal, human…or cell lines.

Of course for cell selection it exists other types of antibiotics, but they are different and their use is more targeted.

When using antibiotics for the first time on a peculiar cell type, it is better to test their toxicity on this cell type.

It depends on your bacterial species and strains. Time to time it should be done.

As has been said that one should master the technique to an extent that there should be no need of any antibiotic in culture work, however if in any case needed, then PEN-STREP-AMPH combinations is common choice. However for selection purposes I use antibiotics like G418 and Hygromycin.

yes, we use antibiotics in tissue culture media but only when we innoculate field grown explants, to stop endogenous microbal infection. Generally we use the antibiotic cefotaxime, we add it in the media itself after autoclaving and cooling it to around 30 degree celsius. Antibiotic addition is not always necessary but it becomes important when we want to control infection

http://www.sigmaaldrich.com/life-science/cell-culture/learning-center/antibiotic-selection.html

Antibiotics Working concentrations and stock solutions.

Hi Yong, 

Thanks for bringing up the important issue. I think its better to improve our working habit/standard rather than follow the substandard procedure. Though I understand the situations in undergrad teaching labs, but it should be kept as a learning curve and follow the experts recommendations in routine culture. 

In my lab, we use 1%pen/strep y.regularl

We do not use antibiotics for maintaining cell lines as antibiotics mask contaminations. However, for experiments we use Pen-Strep and Normocin. However, for antibiotic selections, one must be careful. For example, Pen-Strep can inhibit G418.

We use 1% PSG ( Penicillin-Streptomycin-Glutamine ) regularly

Has anyone intentionally inoculated their cell culture medium with bacteria and fungi to understand what its bactericidal or growth promoting capabilities are? If you understand your risk points then you can tailor your prevention to be based on risk.