270 



Journal of Applied Microscopy. 



Ordinary test-tubes containing glucose bouillon, gelatin, agar, or potato, 

 milk, etc., are inoculated in the usual manner. The projecting part of the 

 cotton plug is cut off close to the mouth of the tube, and the plug is slightly- 

 raised, with sterilized forceps, to facilitate diffusion of the gas. The tubes are then 

 placed in the bottle by means of a pair of rat or crucible forceps and the appara- 

 tus is connected with a Kipp's hydrogen generator. The hydrogen is generated 

 from ordinary granulated zinc by means of commercial sulphuric acid. It is 

 wholly unnecessary to employ chemically pure zinc and acid. The gas should 

 be washed by passage through an alkaline solution of lead acetate and then 

 through a six per cent, solution of potassium permanganate. A rapid current of 

 hydrogen should be passed through the bottle for one to two hours, after which 

 the stopper is turned at right angles. The bottle is then placed in the incubator. 



When the cultures have developed they should be taken out of the bottle 

 and preserved the same as ordinary bacteria. Care should be exercised when 

 removing the stopper so as to avoid breakage. It should first be turned so as 

 to allow air to enter. The thumb and forefinger of the left hand should rest 

 firmly on the shoulder of the stopper while this is gradually worked to and fro 

 with the right hand. This little precaution will prevent the sudden jerking out 

 of the stopper. 



The pyrogallate method can be used in connection with this bottle. Two to 

 three grams of the acid can be placed on the bottom, the inoculated tubes intro- 

 duced, and finally the necessary concentrated alkali ("25 cc. 1:4) can be delivered 

 from a pipette. The stopper must be inserted at once and turned at right angles. 



Fig. 3. 



Fig. 4 



Figs, o and 4 show two forms of apparatus for obtaining plate cultures. The 

 former is provided with a stopper like that in the bottle just described. This 

 apparatus can be used for the gas or the pyrogallate method. The apparatus 

 shown in Fig. 4 is provided with a slightly different stopper, and can be used 

 for the gas, pyrogallate, or vacuum method. 



The lower half of the apparatus (Figs, o and 4) should have an internal 

 diameter of 12 cm. and an inside height of 12 cm. The apparatus can hold six 

 or eight Petri dishes. It can be used for small fiasks or for a large number of 

 tube cultures. The total height of either apparatus should not exceed 24 cm. 

 Each flange should be 2 cm. wide and ;5-4 cm. thick. The outer circumference 



