Why Use Antibiotics in Cell Culture?

Use of antibiotics in cell culture minimizes the loss of valuable cells, reagents, time and efforts due to contamination. Maintenance of aseptic conditions and techniques is vital to a research laboratory that handles cell/tissue culture. Experienced researchers recommend cell culture-tested antibiotics, at adequate concentrations, be used while culturing cells. This practice preserves your cells by preventing contamination that induces morphological or physiological changes. Apart from preventing contamination, certain antibiotics also function as selection agents, used to select and establish transfected/genetically modified cells for research purposes.

• Bacterial and fungal contamination
• Mycoplasma contamination
• Viral contamination
• When should you use Antibiotics in cell culture

Bacterial and fungal contamination

Bacterial and fungal contaminations are most prominent in cell culture because of quick colonization in enriched media. The medium becomes turbid and changes color (pH change) within few hours/days of contamination. Regular microscopic observations and addition of antibiotics to the culture medium prevents contamination from spreading across cultures (see table below).

Antibiotic	Bacteria		Fungi	Yeast	Mycoplasma
	Gram positive	Gram negative
Amphotericin B			✓	✓
Ampicillin	✓	✓
Erythromycin	✓	✓
Gentamycin	✓	✓			✓
Kanamycin	✓	✓			✓
Neomycin	✓	✓
Nystatin			✓	✓
Penicillin-Streptomycin	✓	✓
Polymyxin B		✓
Tetracyclin	✓	✓
Thiabendazole			✓
Tylosin					✓

Select the right antibiotics for your cells.

Determine suitable antibiotic concentration for your cells.

Mycoplasma contamination

Mycoplasma are the smallest, free-living (0.3 µM) prokaryotes observed as filamentous or coccal forms under scanning electron microscope. Mycoplasma contamination is cryptic and extremely challenging to handle. They are undetectable under light microscope but result in morphological changes, chromosome aberrations and altered aminoacid and nucleic acid metabolism. Due to the lack of cell wall they are resistant to routinely used antibiotics. While specific antibiotics are effective, the best agents against mycoplasma are proven to be a combination of multiple biological agents.

The best prevention against mycoplasma contamination is to use cell lines from ECACC that are tested for mycoplasma contamination on regular basis.

Viral contamination

Viruses are most difficult contaminants to detect in cell culture and are difficult to remove. However, the contamination is self-limiting since viruses are solely dependent on host cellular machinery. Virally infected cells pose health hazards to the laboratory personnel. Use of BVDV-tested serum significantly lowers the risk of viruses. 

When should you use Antibiotics in cell culture

• Antibiotics safeguard high value stocks of irreplaceable cultures and to produce working stocks. When reproducible results are critical it is recommended that optimal concentrations of suitable antibiotics be added to cultures.
• Antibiotics like Penicillin-Streptomycin solution at a concentration of 50-100 I.U./ml is recommended for primary cell culture as the chance of contamination is high for the first few weeks. 
• Antibiotics give extra layer of protection from factors like new researchers under training, particulates and aerosols.
  
• Antibiotics are also used to select cells modified by genetic engineering (selection agents). Puromycin and hygromycin B are used to select and establish cells expressing the puromycin-resistance and hygromycin-resistance genes, respectively.

It is wrong to assume that antibiotics are entirely bad for cell culture; the correct approach is to use the optimum concentration in your cell culture experiments to prevent contamination and protect cells in your lab.

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