INOCULATION OF TUBES 87 



upon the side of the tube at the margin of the fluid, and 

 the emulsion washed down by tilting the tube. The 

 inoculation having been completed, the plug is quickly 

 replaced, and the needle is again heated in the flame to 

 destroy the remains of the growth upon it ; sterilisation of 

 the .needle before and after use must become a habit and 

 must never be omitted. If the original culture is in a deep 

 stab or a fluid medium, a looped platinum needle may 

 sometimes be used with advantage. For greater safety 

 the plugs, before being replaced, may be singed in the 

 flame, and pushed while burning into the tube, and this 

 procedure must always be adopted if the plug be dropped 

 or brush against anything. If the tubes have to be kept 

 for any length of time, especially in the blood-heat incu- 

 bator, each should be capped with a rubber cap, tinfoil, 

 or gutta-percha tissue which has been soaked in 1-500 

 corrosive sublimate solution. 



Anaerobic cultures. Many organisms will not grow in 

 the presence of free oxygen, and various expedients have 

 to be adopted to exclude or remove the air. Removal of 

 air with an exhaust pump may be employed, but is in- 

 convenient and not often used. The simplest method is 

 to make the cultivation in a deep stab in glucose-agar or 

 gelatin. Narrow test-tubes filled three parts full with the 

 medium are best, and immediately before use they should 

 be boiled in water for five minutes, and then solidified in 

 cold water. The object of this is to soften the medium so 

 that it does not split, as a dry medium will, when the 

 needle is plunged into it ; moreover, the needle track 

 closes up more readily, and the dissolved oxygen is 

 expelled. When the tubes are solid, each is inoculated by 

 a single steady puncture down the middle, the wire being 

 straight or with a closed loop at the end. After inocula- 

 tion, each tube is carefully heated at the upper border 

 of the medium so as to melt this slightly and seal the 



