126 MANUAL OF MICROBIOLOGICAL METHODS 



Method. Place small amount of tap water in bottom of cylinder to 

 remove the P2O5 which forms. Stack inoculated plates or tubes on sup- 

 port. Add tube of methylene blue solution (see page 122). Place smal) 

 (50-ml) beaker on top of cultures. Remove two or three short (l}i- to 

 2-in.) pieces of phosphorus from water with forceps or tongs, and place in 

 beaker. Immediately put lid on jar and seal with plasticene. (Upon 

 drying for a few minutes the phosphorus should ignite spontaneously and 

 remain burning as long as there is oxygen present.) If experience shows 

 that the phosphorus used does not ignite spontaneously but merely gives 

 off a gray smoke, ignite it before the jar is sealed by a match held with the 

 forceps. Since considerable heat is developed, place beaker, unless resist- 

 ant glass is used, 3 in. from the top of the container and put a '^ blank'' 

 plate under the beaker rather than an inoculated plate. After the phos- 

 phorus ignites and the jar is tightly sealed, place it directly in the incu- 

 bator. At the time the container is opened have available a crock or pan 

 filled with water. As soon as the lid is taken from the jar, remove the 

 beaker containing the phosphorus with the tongs and submerge under the 

 water in the pan and save for later use. After this remove the cultures 

 from the jar. 



Advantages. Quick method of obtaining anaerobiosis. It is relatively inexpensive 

 since the only materials are phosphorus and a container which may be sealed. Dis- 

 advantages. Care must be exercised to prevent accidental burns which are very pain- 

 ful. Inexperienced technicians should be cautioned concerning the dangers. 



Alkaline Pyrogallol Methods 



Another chemical method for removing oxygen in order to promote 

 anaerobic growth is to utilize the oxygen absorptive capacity of the 

 reaction between alkali and pyrogallic acid. Of the technics and devices 

 reported which make use of this reaction two may be recommended as 

 being especially useful. One of these concerns a technic applied to indi- 

 vidual plate culture, and the other relates to a system for individual tube 

 cultures. 



Spray (or Bray) Plate Cultures 



Materials. (1) Spray (1930) anaerobic dish;^ (2) plasticene, or tape 

 for sealing; (3) 20 per cent aqueous NaOH; (4) 40 per cent aqueous 

 pyrogallic acid. 



Note: The Spray dish consists of an ordinary glass petri-dish top and a special 

 bottom which is deep and has a raised ridge across the center. The top of the bottom 

 dish has a lip into which the top section of the dish fits. Although constructed of 

 heat-resistant glass, in practice considerable breakage may be encountered during 



^ Spray anaerobic dish. Fisher Scientific Company, Pittsburgh, Pennsylvania, or 

 E. H. Sargent Company, Chicago, Illinois, 



