MECHANICAL METHODS 81 



of the moist chamber or with the glass stands, and this explains why perchloride 

 can be used for sterilizing these pieces of the apparatus. 



7. In the same way pour a plate with tube No. 2, place it in the moist chamber, 

 and put the third glass support in position. 



8. Repeat the process with tube No. 1, and place it on the third plate-rest. 



9. Put the moist chamber in the incubator at 20 C. The plates have been arranged 

 so that the one containing the largest number of organisms is nearest the top of the 

 chamber. Colonies will appear on it earlier than on the other plates, and it can be 

 examined and studied without touching the latter, which should not be interfered 

 with until growth appears on them. 



C. Esmarch's tubes. 



Apparatus required. 1. Three Pasteur pipettes. 



2. Three tubes of gelatin. The tubes should be rather longer and wider 

 than ordinary culture-tubes, and each should contain 10 c.c. of sterile gelatin. 



3. Three sterile india-rubber caps. 



Technique. 1. Sow the tubes 1, 2, and 3 as before (p. 78 A (i)). 



2. Slip an india-rubber cap over the wool plug of each. 



3. Cool each tube in turn under the cold water tap : hold it as nearly 

 horizontally as possible, so that the gelatin coats the whole of the inner 

 surface of 'the tube below the plug (but without touching the wool), and 

 rotate it between the index finger and thumb of each hand. When the 

 gelatin sets it is thus spread in a thin layer over the whole of the inner surface 

 of the. tube and forms a " roll " tube. 



4. When the gelatin has set, take off the india-rubber cap and incubate 

 the tubes at 20 C. 



This method has the great advantage of absolutely preventing any contamination 

 of the medium, but the investigation of the colonies which develop is. rendered 

 more difficult by the cylindrical shape of the gelatin surface. 



2. Dissemination on the surface of a solid medium. 



When it is necessary to isolate the organisms present in a non-liquid pro- 

 duct such as a false membrane, viscous sputum, etc., a small portion of the 

 material is smeared over the surface of some solid medium contained in a 

 Petri dish or sloped in a tube. This, which is the method now universally 

 adopted for the isolation of diphtheria bacilli from false membranes, is 

 available when the media which it is proposed to use cannot be liquefied 

 by heat, e.g. potato or serum. 



If there be reason to suppose that the material 

 under investigation is very rich in organisms 

 (excreta, for example), a small portion is diluted 

 in a few cubic centimetres of broth or sterile 

 water and a trace of the dilution used for 

 sowing cultures.- 



Two methods are available. 



A. Stroke cultures. 



The method of isolation on agar plates will 

 be taken as an illustration (fig. 73). 

 Apparatus required. 1. A medium or stout 



i , FIG. 73. Isolation of organisms 



platinum Wire. by parallel stroke culture on Petri 



2. A tube of agar. disheg . x i 



3. A sterile Petri dish. 



Technique. 1. Melt the agar and pour it with the usual precautions into 

 the Petri dish. 



