' M. H. SOULE 253 



sure is likely to exceed the capacity of the U-tube; and (2) to enable one to withdraw gas 

 from the culture tube without sucking over the mercury column. Likewise, when evacuating 

 the culture tube with the object of refilling it with fresh air, or with O2, CO2, etc., this cock 

 must be closed. At all other times it is kept open. 



Stopcocks I and 3 are used to equilibrate the mercury levels at the beginning of a test, 

 after which both are closed. By means of tailcock 3 a sample of the gas content in the cul- 

 ture tube or jar is easily withdrawn for analysis. It is also the means by which the gas within 

 the culture apparatus is evacuated and replaced with fresh air, or with any desired gas. 



At this place it may be well to indicate the ease with which desired gas tensions may be 

 approximated by the use of an evacuating apparatus which consists of a Chapman water 

 pump, a vertical manometer, and a Woulff bottle. One tubulure of the latter is closed with a 

 rubber stopper through which is inserted one of the main arms of a three-way cock. The 

 midarm of this cock, horizontal in position, is joined to a glass Y-shaped connector with cocks 

 on each arm. The upper main arm is attached to the descending limb of a vertical mano- 

 meter. Another tubulure is connected with the pump by means of a three-way tailcock. This 

 serves to disconnect the latter when the desired negative pressure has been obtained. 



If it is desired to introduce into a jar a certain percentage of CO2, for example, 40 per cent, 

 this can be readily done. Assuming the barometer to read 750, and the thermometer 22.3°, 

 the corresponding aqueous tension is 20 mm. of mercury. Hence 5 — / = 750— 20 = 730 

 mm. and 40 per cent of this = 292. This value represents the partial tension of the gas to be 

 admitted, provided it is dry. Connection is made to the jar by way of one arm of the Y; the 

 cock on the other arm is closed. The pump is started and evacuation continued until the 

 manometer registers 292 mm. The three-way cock is now turned so that the connection to 

 the Woulff bottle is closed, and pure CO2 (from a tank or generator)' is run in through the 

 other arm of the Y-connector until the manometer reads zero. By means of the foregoing 

 procedure, it is possible to introduce into the jar any desired amount of a given gas or various 

 mixtures of gas, and with care the desired tensions may be easily obtained within one-half 

 of I per cent. For exact work a sample of the gas should always be withdrawn and analyzed. 

 Satisfactory manometric readings may be obtained with culture tubes, but they are not suit- 

 ed for the withdrawal of 10-20 cc. of gas for analysis. 



There is another important consideration, viz., the necessity of providing for a sufficient 

 volume of O2 to permit the full growth of the organisms; for example, it was found that the 

 human variety of the tubercle bacillus required at least 500 cc. of air to produce a rich growth 

 in a tube. Hence if the //-tube, with a volume of 100 cc, is used, little or no visible growth 

 will occur. For this reason it is preferable to make use of the large chamber or jar (Fig. 2) of 

 known volume which can easily be made gas tight. These jars can be obtained in two sizes 

 which have approximate air capacities of 2,400 and 3,400 cc, respectively. 



Culture tubes, flasks, or Petri plates may be placed in the jar which can then be used with 

 air or filled with any desired gas. In order to develop quickly the full aqueous tension in the 

 container it is necessary to place a few drops of distilled water on the floor of the jar. 



Rubber tubing is not desirable for connecting the respiratory chamber to a manometer. 

 It is well known that CO2 diffuses through a rubber tube, and without doubt it would be 

 preferable to have all-glass connections. However, a satisfactory connection can be made by 

 bringing the end of the manometer into contact with the glass stopper of the respiratory 

 chamber by means of the special No. 25 rubber stopper (Fig. 2). Negative or positive pres- 

 sures of 200 mm. or more have held for weeks. 



The gases may be analyzed by any one of the many standard methods. The modified 



' Gases procured in ordinary commercial tanks must be purified before use or the percentage of 

 foreign substances determined and allowed for in the calculations. 



