ISOLATION OF BACTERIA 41 



sufficient to keep the gelatin liquid, but not high enough to 

 destroy the vitality of the bacteria which are to be the 

 subjects of experiment. The tubes are then numbered 

 1, 2, and 3. By means of a platinum needle, which 

 has been previously sterilised at red heat and allowed 

 to cool, after carefully withdrawing the plug, a mere trace 

 of the mixture of organisms under examination is intro- 

 duced into tube 1 and well mixed. If the material is 

 too coherent, attempts must be made to separate the 

 organisms by rubbing them with the point of the platinum 

 wire against the side of the tube below the surface of the 

 gelatin. The plug is replaced and the needle sterilised 

 by passing it through the flame. A platinum loop is 

 sterilised, and whan cool, a loopful of the gelatin transferred 

 from tube 1 to tube 2, and the contents well mixed, after 

 which two or three loopfuls from tube 2 are transferred 

 to tube 3. By the above dilution the number of organisms 

 in the third tube is probably so small as to give a satis- 

 factory ' plate.' It may happen that the dilution is 

 carried too far, in which event the .plate required is ob- 

 tained from the second tube. Success in this operation 

 is a matter of experience and judgment. When inocula- 

 ting tubes the cotton- wool plugs must be held between the 

 fingers, best between the third and fourth, using the back 

 of the hand, and must be carefully returned into the tube 

 after inoculation, without the part that goes in the tube 

 being allowed to come into contact with the surface of 

 the hand or bench. The manner in which plugs and 

 tubes are held is largely a matter of taste, but whatever 

 posture is adopted, the positions of the respective 

 plugs and tubes must be remembered, to avoid mixing 

 them. 



The plug of No. 1 tube is flamed and removed, the lips 

 of the tube are flamed, and the contents of the tube poured 

 into a Petri dish which is marked ' No. 1.' The tubes 

 marked 2 and 3 are treated similarly. Petri dishes are 

 from 10 to 20 centimetres in diameter and about 1-5 or 2 

 centimetres deep, and have loosely fitting covers of the 

 same form as the dishes themselves. The colonies may 

 be examined and counted without removing the lid. 



Esmarch's Roll Cultures. The inoculated liquefied 

 gelatin is distributed in a thin layer upon the walls of a 

 wide test-tube by rotating the tube upon a block of ice 



