Preparation of Stock Solution of L-Glutamine (200 mM)


  • L-Glutamine (C5H10N2O3; CAS number: 56-85-9) is an important component of the cell culture medium. It is an alternative energy source especially for the cell types with high metabolic demands (e.g., fast-growing cell lines) and consumes glucose inefficiently. In addition, it is also a source of nitrogen and carbon for the synthesis of cell components including nucleic acid and proteins.
  • It is unstable in solution, therefore it is recommended to add L-glutamine in basal medium just prior to use. The L-Glutamine supplemented medium can be stored at 4 °C just for 2 weeks.
  • The concentration of L-Glutamine may vary depending on cell types and culture medium. However, for most classical serum-grown cell lines, a 2 mM concentration is generally used. 
  • As compared to other amino acids, a relatively high concentration of L-glutamine is used in cell culture medium (vary from 2 to 10 mM) as it is a growth-limiting amino acid for the growth of most cell types.
  • It is soluble in water (solubility in water at 25 °C: 36 mg/ml) and yields a clear colorless solution.
  • It degrades in the culture medium that results the accumulation of toxic of ammona in the culture medium.
  • Since autoclaving degrades the glutamine solution, filter sterilization (filtering through a sterile membrane with pore size 0.22 micron or less) is used to sterilize L-glutamine solution.


> L-Glutamine (Molecular weight: 146.1456)

> Cell culture grade water (Milli-Q water) or 0.85% saline solution

Equipment and disposables
> Conical flask / Beaker (capasity 250 ml)

> Measuring cylinder
> Magnetic stirrer/glass rod

200 mM L-Glutamine in water or 0.85% saline

Preparation of 100 ml of 200 mM L-Glutamine solution in water or 0.85% saline solution
Note: Glutamine solution can be prepared either in water or in 0.85% saline.


Step 1: To prepare 100 ml of 200 mM L-Glutamine solution, dissolve 2.92 g L-Glutamine in 90 ml cell culture grade water or 0.85% saline.
> Weigh out 2.92 g of Glutamine and transfer it to a container (beaker/conical flask, capacity 250 ml) containing 90 ml water or 0.85% saline.
> Stir the solution using a glass rod/magnetic stirrer until all L-Glutamine dissolves completely.

> Capacity of the container (beaker/conical flask) must be at least double the volume of the solution you intended to prepare. It helps to prepare the solution without spilling the solution out of the container.

> Make sure no undissolved L-Glutamine in the beaker.

Step 2: Adjust the volume to 100 ml with the solvent you used for the solution preparation (cell culture grade water or 0.85% saline).
> Transfer the solution to a volumetric flask or measuring cylinder. Adjust the final volume of 100 ml with the solvent. 
> After adjusting the volume 100 ml, transfer the solution back to the same container.

Step 3: Filter sterilize the solution
> Sterilize the solution by filtering it through a sterilized filter unit containing a filter paper with pore size 0.22 µm or less.

The solution can not be autoclaved as autoclaving will destroy the L-Glutamine.

Store the solution in small aliquots (5 ml) in the freezer (-20 °C). The solution is stable at -20C for several months.

Use Calculator to calculate the quantities of NaOH amount for the preparation of NaOH solution of specific concentration and volume

Molecular weight of NaOH: 40 g/mole

Volume of Sodium hydroxide solution: ml

(Change the volume of the solution)

Molarity of Sodium hydroxide solution: M

(Change the molarity of the solution)

Amount of sodium hydroxide: 40 g

Was this post helpful?

Author: admin

Leave a Reply

Your email address will not be published. Required fields are marked *