CH. iv.] PREPARATION OF CULTURE-MEDIA. 33 



their cotton-wool plugs, before receiving the material, should 

 be thoroughly sterilised by overheating. As I mentioned in 

 the previous chapter, this ought to be well borne in mind, for 

 starting with a sterile nourishing fluid i.e. one that has been 

 kept in the stock flask for several days to several weeks in 

 the incubator at a temperature of from 32 to 38 C. and that 

 has remained perfectly clear and limpid and working with 

 thoroughly sterilised test-tubes and cotton-wool plugs very 

 little care is required to obtain sterile material ready for 

 inoculation. To start with a stock of nourishing material, 

 however well sterilised, and to decant it into test-tubes with 

 cotton-wool plugs not absolutely sterile must lead to failure. 

 I have seen this happen over and over again, and all the 

 material decanted became consequently contaminated and 

 thereby useless for inoculations. The test tubes and flasks 

 must be well cleaned, then dried, placed in the air-chamber, 

 and kept there exposed for several hours to a temperature of 

 from 130 to 150 C. on several successive days, or they 

 may be thoroughly heated in all parts over the open flame 

 of a gas-burner. The same applies to the cotton-wool, as 

 mentioned in a former chapter. The test-tubes and flasks 

 are plugged by means of clean forceps with the cotton-wool 

 which is just brown, and then replaced in the air-chamber 

 and again heated for several hours on two or three occasions 

 up to a temperature of 130 to 150 C., or they may be well 

 heated over the open flame of the burner. To decant sterile 

 stock fluid into these test-tubes and flasks I proceed thus : 

 A clean beaker with spout, covered with a clean glass plate, 

 is placed on a wire net on a tripod over the flame of a 

 Bunsen burner, and thoroughly heated for half an hour or 

 so ; then it is allowed to cool, and when cool, the plug of the 

 stock flask is lifted with forceps, and some of the sterile 

 fluid quickly poured from the flask into the beaker. The 



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