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How to Purge Cell Culture Medium with Nitrogen

Biological studies use cell cultures to investigate the effects of drugs, therapies and practices on the life cycles and activity of cells. The growth of cells and maintenance of cell lines occur in culture media that is beneficial to the cells in question. The media provides all the nutrients and environmental conditions required by the cells. Most cells need a source of nitrogen to remain healthy and to supply some of the requirements for the many processes the cells undergo during their life cycle. While nitrogen is obtainable from ammonia and other nitrogen-containing substances, purging the media also provides a source for the needed nitrogen.

Things You'll Need

Cell culture medium
Large beaker
Mechanical stirrer
Nitrogen tank
Sparging tube

Instructions

- 1
  Pour your cell culture medium into a large beaker and place the beaker on a mechanical stirrer. The stirrer mixes the medium during purging by the nitrogen stream. Using this method will allow a preparation of a large amount of medium to maintain more uniform composition.
- 2
  Affix a sparging tube to the output of a nitrogen tank. A sparging tube is composed of a glass tube with a scintered glass tip incorporated into the end of the tube. The open end of the tube attached to a length of plastic tubing connected to a nitrogen tank provides a flow of nitrogen as it exits the scintered glass end of the tube by many streams of tiny gas bubbles.
- 3
  Place the scintered glass end of the sparging tube into the bottom of the beaker and turn on the mixer. Set the stir rate of the mechanical stirrer to provide even mixing of the medium.
- 4
  Turn on the flow of nitrogen at the tank and allow it to bubble into the medium. The nitrogen will saturate the medium with nitrogen as it dissolves. Each liquid possesses a finite solubility of different gases. As the nitrogen dissolves in the medium, other dissolved gases are expelled from the medium. Purging removes gases such as oxygen and carbon dioxide from the culture medium.
- 5
  Allow the nitrogen stream to bubble at least 15 minutes to saturate the medium, depending on the flow rate of the gas. Once the purging of the medium is finished, remove the scintered glass tube from the beaker and transfer the purged cell culture medium to a closed container and store it in the refrigerator. The reduced temperature of the refrigerator will reduce the degree at which gasses dissolve into the medium during storage.

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