Glutamine is a non-essential amino acid for the body, however mammalian cells in culture require the presence of millimolar concentrations of this amino acid whose presence is essential for cell growth and proliferation, especially in cancer cells. Glutamine takes part in numerous intracellular metabolic reactions both as a nitrogen and carbon donor (Zhang J et al., 2017).
All cell culture media must therefore contain this amino acid.
The concentration of glutamine required in the culture medium varies according to the cell lines in culture. In most cases a 2 mM concentration is used
Gln or not in my purchased cell culture medium?
If available, it is best to avoid purchasing culture media already complemented with glutamine, or it is better to choose a medium supplemented with L-glutamine in dipeptide forms, such as alanyl-l-glutamine and glycyl-l-glutamine.
In fact, glutamine degrades rapidly and non-enzymatically in solution, with the formation of pyrrolidonecarboxylic acid and ammonia toxic to cells.
Due to this instability in solution it is generally suggested to add glutamine to the culture medium just before its use. After addition, the medium can be kept in the refrigerator for a maximum of 2-3 weeks.
Several culture media already pre-completed with glutamine have a lower than nominal glutamine concentration and a certain ammonia concentration that increases over time upon their arrival in the laboratory.
This occurs at both 20 ° C and 4 ° C ( Heeneman et al., 1993).
Serum concentration, on the other hand, has little or no effect on the rate of glutamine decomposition (Ozturk et al. 1990).
Ammonia is mainly present in the form of NH4+ cation at physiological pH.
While in vivo ammonia is metabolized to urea and eliminated, in vivo it accumulates as NH4+ in the medium, with toxic effects for the cells.
Preparation of a 200 mM glutamine stock solution.
Although commercially available, a 200 mM mother of glutamine can be prepared quickly and easily in the laboratory, with good savings.
- Weigh 2.92 g of glutamine powder and put it in a beaker.
- Add containing 90 ml PBS.
- Stir the solution using a magnetic stirrer until all L-Glutamine dissolves completely.
- With the use of a graduated cilinder, adjust the final volume of 100 ml with PBS.
- Sterilize the glutamine solution by filtering it through a sterilized filter unit with pore size 0.22 µm or less.
DO NOT autoclave! The high temperature degrades glutamine
Aliquot in disposable aliquots (avoids freezing and thawing cycles). In -20 the solution is stable for 6 months