GasPak Anaerobic System: Principle, Application

GasPak anaerobic system is used to create an oxygen-free environment for the growth of anaerobic microorganisms. At present, it is the most commonly used method for anaerobiosis. It is very simple to perform and is perfect for a laboratory having less sample load.

Components of GasPak Anaerobic System

1. polycarbonate jar (or anaerobic container)
2. a lid with a gasket to prevent airflow (for jar system)
3. Indicator strip (a strip impregnated with an oxidation-reduction indicator such as methylene blue or resazurin)
4. disposable gas generating pouch (a pouch containing sodium borohydride and sodium bicarbonate)
5. a palladium catalyst

Principle

Inoculated plates or tubes are placed inside the polycarbonate jar or anaerobic container along with gas generator sachet and indicator strip and is sealed completely. In the presence of water, chemicals present inside the sachet i.e. sodium bicarbonate (NaHCO₃) and sodium borohydride (NaBH₄) react chemically producing hydrogen and carbon dioxide gas.

The hydrogen thus produced reacts with oxygen present inside the jar producing water (which forms as condensation on the inside of the jar).

2H₂ + O₂ + catalyst = 2H₂O

This reaction is catalyzed by the element palladium, which is attached to the underside of the lid of the jar. The carbon dioxide replaces the removed oxygen, creating a completely anaerobic environment.

Note: Current anaerobic gas-generating systems do not require a catalyst and/or the addition of water to activate these systems. Please refer to the user manual provided by the supplier before commencing your works.

Steps

1. Inoculate agar plate with test strain.
2. Place inoculated agar plates in an anaerobic jar, container, or pouch.
3. Add anaerobic indicator strip.
4. Add the required number of gas generator sachet after opening the sealed GasPak container and place the lid in place or seal the pouch/container.
5. Incubate at the desired temperature.
6. Check anaerobic indicator strip for color change.

Note: These steps may slightly vary based on commercial suppliers. Please refer to the user manual provided by the supplier.

Indicator of anaerobiosis

The effectiveness of GasPak anaerobic system can be checked by the following methods:

1. Anaerobic indicator strips:
   1. Impregnated with methylene blue: remains colorless in anaerobic conditions, but turns blue on exposure to oxygen.
   2. Impregnated with resazurin, a redox indicator: colorless (white) in anaerobic conditions but turns to pink in the presence of oxygen.
2. Biological indicator: A plate inoculated with Pseudomonas is incubated along with other inoculated plates for anaerobic cultures. The absence of growth of Pseudomonas (which is an obligate aerobe) indicates that perfect anaerobiosis has been achieved.

Commercial products

BD GasPak EZ gas generating system is the first complete, waterless and catalyst-free system that produces anaerobic, microaerophilic, or carbon dioxide enriched environment. This fully integrated system is easy to use and is available on two systems; container system & pouch system. BD container system is available in 10, 15, and 30 Petri dish capacity. Pouch system is available in 1-4 petri dish capacity with all needed materials (pouches, sachets, and indicators).

References and further reading

• Koneman’s Color Atlas and Textbook of Diagnostic Microbiology
• BD GasPak systems