When is the best time to add antibiotics in cell culture?

[Goodie]

should i add antibiotica :

1. in medium after thawing?
2. in freezing medium?
3. in incubation of transfected cells?

or should i add antibiotica only when my mammalian cells have a vector with the appropriate selectable marker for that antibiotica?

thanks.

 

[iansmith]

The sterile antibiotic should be added to the medium just prior to inoculation and only when your cell carry the selective marker.

 

[DocToxyn]

There are numerous cell lines that are routinely kept in media that include antibiotics such as penicillin, streptomycin, gentamicin, etc., some even add antimycotics such as amphotericin. These aren't there for any selection purposes, rather they are there to keep the cultures free of bacteria and other nasty things that will out compete your cells. These media additions and their concentrations are usually found in the methods sections of papers that use the cell line you are interested in. If the cells you have are commonly cultured with these agents, then its best to use them since the cells are "used to" living with them and changing this might alter their function/responses. They are however, not an excuse for poor culture handling practices and should not be entirely relied upon to maintain clean cultures.

As far as freezing medium, many use the standard culture medium containing all components in addition to ammendments like higher serum levels and/or dimethyl sulfoxide (DMSO, ~10-20%) to reduce freezing induced damage.