APPENDIX l6l 



I litre. Add 120 grammes of best "gold label" gelatine (i.e. 12 

 per cent.). Place the flask in the steam sterilizer and steam for 

 one hour to completely dissolve the gelatine. Estimate the 

 reaction and add the calculated amount of alkali to bring to 

 a + ro per cent, reaction. The increased bulk of the gelatine 

 brings the contents up to 1096 c.c. (Eyre) so the amount of 



X 



- alkali which has to be added must be estimated on this basis, 

 o 



Cool down to about 50 60 C. and add the white of one 

 egg previously mixed with a little water. Mix well and keep 

 in current steam for about 30 40 minutes. The egg albumin 

 is coagulated and acts as a mechanical clarifying agent. The 

 gelatine above the clots should be clear. Filter into a clean 

 flask, funnel and flask being placed in the steam sterilizer. Put 

 into tubes (which should have been previously sterilized), and 

 sterilize for thirty minutes on three successive days. 



Sometimes considerable difficulty is experienced in obtaining 

 gelatine media sterile. If this occurs great care should be taken 

 to have all flasks, test-tubes, cotton-wool, and other articles 

 used in the manufacture previously sterilized. It must be 

 remembered that gelatine media must never be heated above 

 1 00 C. and should not be heated more than is necessary. For 

 this and all media it is important to test the sterility of the 

 medium by incubating the finished media for several days at 

 37 C. 



Nutrient Agar. Weigh out 5 grammes Lemco, 5 grammes 

 sodium chloride, 10 grammes peptone (Witte's) and heat to 

 boiling with a litre of distilled or rain water in an enamelled 

 saucepan. The solution must be distinctly alkaline. Transfer 

 to a flask and make up to one litre. Add 15 grammes (i.e. 

 I '5 per cent.) of thread or powdered agar. Digest in the auto- 

 clave at 115 C. for forty-five minutes. Estimate the reaction 

 and bring to a + I per cent, reaction. Cool to about 50 60 C. 

 and add the white of an egg previously mixed with a little 

 distilled water. 



Heat in autoclave at H5C. for forty-five minutes. Filter, 

 preferably through papier chardin, flask and funnel being placed 



s. w. ii 



