BACTERIOLOGICAL TECHNIC 



67 



cock, and glass nib attachment. Occasionally it is desirable to place 

 exact amounts of culture media in the tubes, in which case a graduate, a 

 burette, a pipette, or other convenient measuring device may be used. 



4. Sterilization of Culture Media 



All culture media in tubes as above set forth, and the portions remain- 

 ing after the desired number of tubes are filled, must be considered as 

 being contaminated with living microbes and their spores. These mi- 

 crobes and spores are killed by the sterilizing 

 process. " For all ordinary purposes the dis- 

 continuous, or fractional method answers the 

 purpose admirably. Place the test-tubes, 

 flasks, and other cotton-plugged containers 

 with culture media, in a steam sterilizer 

 (Arnold steam sterilizer, either board of health 

 or cylindrical form ; or kitchen vegetable cooker 

 or steamer). The test-tubes are placed in 

 wire baskets (rectangular or cyhndrical). 

 These several containers with culture media 

 are exposed to live steam for about thirty 

 minutes, whereupon the flame is turned out, 

 and if convenient the containers are allowed 

 to remain in the sterilizer. Caution must be 

 observed to guard against condensed steam 

 running into the several containers. The 

 better way is to remove the containers and 

 place them in an incubator kept at a tem- 

 perature of 20° C. In twenty-four hours, or 

 thereabouts, steam is again applied for thirty 

 m'nutes. This is repeated a third time on 

 the second day after the first sterilization. 

 The first sterilization , presumably kills most of 

 the vegetative cells. During the first interval 

 of twenty-four hours most of the spores present 

 develop into vegetative ceUs, which are killed 

 at the second sterilization. Should any sur- 

 vive the second steaming, they are sure to be 

 killed during the third sterilization. During 

 this time the cotton plugs have not been 



removed. The media thus fractionally or discontinuously sterihzed are 

 now ready for use in making microbic cultures, or they may be set aside 

 for an indefinite period of time. 



Fig. 16. — Autoclave for 

 using steam under pressure 

 for purposes of sterilization. 

 Many different forms and sizes 

 of autoclaves are on the 

 market. Some of them to be 

 used with gas heat, others 

 with electricity. The enor- 

 mous autoclaves used by the 

 large canneries will hold sev- 

 eral tons. Enormous sterili- 

 zers on the order of the auto- 

 clave are used at the national 

 quarantine stations, as at 

 Angel Island, San Francisco, 

 and at New Orleans. 



