McIntosh and Fildes’ anaerobic jar is an instrument used in microbiology laboratory for the generation of anaerobic conditions (anaerobiosis) to culture obligates anaerobes such as Clostridium spp. Anaerobiosis obtained by McIntosh and Fildes’ anaerobic jar is one of the excellent and most widely used methods for anaerobiosis but it requires costly special apparatus and a vacuum pump. Availability of gas supply is another major drawback of this system. Currently, it is being replaced by a more convenient GasPak system.
About the equipment
McIntosh and Fildes’ jar consists of a 8*5 inch (20*12.5 cm) jar of stout glass or metal with a tight-fitting metal lid. The lid can be clamped airtight with a screw and is fitted with two tubes with taps, one for the introduction of the gas inside (inlet) and the other as an outlet for the vacuum valve. The lid also contains two terminals that can be connected to an electric supply. A capsule containing alumina pellets coated with palladium (palladinised alumina) is suspended under the lid by stout wires which are connected with the terminals to heat the catalyst for its activity. Nowadays, catalysts that are active at room temperature are also available.
McIntosh and Fildes’ anaerobic jar works on the principle of evacuation and replacement, where the air inside the chamber is evacuated and replaced with mixture of gases (consisting of 5% CO2, 10% H2, and 85% N2).
It is practically impossible to evacuate all the air so some amount of oxygen will still be left behind. The residual oxygen left behind is converted to water using spongy palladium or platinum catalyst. The catalyst acts as a catalyzing agent causing slow combination of hydrogen and oxygen to form water. Reduced methylene blue is generally used as an indicator (mixture of NaOH, methylene blue, and glucose). It becomes colorless anaerobically but regains blue color on exposure to oxygen.
- Keep the inoculated culture plates inside the jar along with an indicator.
- Screw tight the lid
- Close the inlet tube and connect outlet tube to a vacuum pump (at least three quarters of the air of the jar can be removed).
- Note the pressure on a vacuum gauze and when the pressure is reduced to 100 mm Hg (i.e., 600 mm below atmospheric), tightly close the outlet tap.
- Connect the inlet tap is to a hydrogen supply and then open it. Hydrogen is passed through a small wash bottle.
- Bring the reduced pressure up to 760 mm Hg (i.e., atmospheric) by monitoring the vacuum gauze as 0.
- Switch on the electric terminals for heating the palladinised crystal (When room temperature catalyst is used heating is not required).
- The catalyst helps the combination of hydrogen and residual oxygen to form water. This process is allowed to continue for 20 minutes.
- Incubate the McIntosh and Fildes’ jar in an incubator at 37°C for 48 hours.
Monitoring efficacy of anaerobiosis:
Reduced methylene blue indicator is used to check the efficacy of anaerobiosis. A tube containing reduced methylene blue solution had to kept inside the jar along with the culture plates. Methylene blue is colorless in reduced conditions and turns blue when oxidized.