11143-12 MANUAL OF METHODS FOR PURE CULTURE STUDY 



large milled head sufficiently to make the lid gas-tight but not to the 

 point at which the action of the coiled spring is ineffective. Tighten 

 the lock nut (the smaller and concentric milled head). Introduce 

 hydrogen from cylinder, through reducing valve set for two pounds, 

 and keep flowing for two minutes or more. Test whether or not 

 all the air has been removed by attaching a rubber hose to the exit 

 valve and allowing the gas to excape in a cup of soapy water. If 

 the gas bubbles fail to "explode" when a lighted match is applied 

 but ignite to burn with a non-luminous flame, the concentration of 

 hydrogen is sufficient to proceed. Close both valves and connect 

 the wiring terminals to an electric source of correct voltage and 

 through a 0.6-0.7 ampere resistance. Formation of droplets of water 

 on the inside walls of the jar indicates correct functioning of the 

 apparatus. After a negative pressure develops (a few minutes) 

 add more hydrogen slowly. Continue the current for 30 minutes. 

 Then tighten the valves of the jar and remove the electric connection. 



Advantages and disadvantages: See above for Brewer jar. Apparently there is 

 greater danger of explosions with the Mcintosh and Fildes jar than with the Brewer 

 jar. Inexperienced technicians are warned to proceed with caution when using this 

 apparatus. 



Plating System Using Strongly Reducing Medium 

 Recently there has been introduced by Brewer (1942) another 

 single plating device which has much to recommend it. Because 

 of its promise it is introduced here even though it has not as yet been 

 used sufficiently widely to establish a reputation. The dish must 

 be used with an agar containing highly reducing agents. The design 

 of the dish is such that the top of the dish rests, at its periphery, 

 on the medium to form a seal, and the remainder of the dish is slightly 

 raised. Thus only a small amount of air is trapped over the surface 

 of the agar and this is removed by means of the reducing action of 

 the medium. 



brewer culture dish'^ 



Materials: (1) Brewer anaerobic culture dish; (2) regular petri 

 dish with bottom either 15 mm. or 10 mm. deep; (3) infusion agar 

 suitable for anaerobes which contains suitable reducing agents, such 

 as the following: 0.2% sodium thioglycoUate, 0.1% sodium form- 

 aldehyde sufoxylate, and 0.0002% methylene blue. 



Method: Pour sterilized medium in bottom of regular petri dish 

 (25 ml. minimum in 10 mm. dish, and 40 ml. minimum in 15 mm. 

 dish). Streak center area from sample or culture. Replace the 

 lid of the regular dish with the Brewer anaerobic lid. (The lid 

 at its periphery, should touch the agar at all points in order that 

 a perfect seal be obtained. In the successfully prepared dish, the 

 agar in the center of the dish remains colorless while the blue color 

 returns to the agar at the edge of the dish due to oxygenation of the 



^Brewer anaerobic dish. Baltimore Biological Laboratory, Baltimore, Md., and 

 Kimble Glass Company, Vineland, New Jersey. 



