46 METHODS OF CULTIVATION OF BACTERIA. 



reached. Repeat process thrice and take mean to obtain 

 amount of soda required. From this calculate acidity of 

 medium per litre, and neutralise with normal soda solution. 

 Check calculation by a fresh titration of 5 c.c. of the 

 neutralised medium. Steam for half an hour, and take 

 reaction again to see that it is constant. Now add normal 

 HC1 in the ratio of 1.5 c.c. per cent. 



The gelatine and agar media (vide infra] are treated in 

 the same way. 



i (b). Glucose Broth. To the other constituents of 

 i (a) there is added i or 2 per cent of grape sugar. The 

 steps in the preparation are the same. Glucose being a 

 reducing agent, no free oxygen can exist in a medium 

 containing it, and therefore glucose broth is used as a 

 culture fluid for anaerobic organisms. 



1 (<;). Glycerine Broth. The initial steps are the same 

 as in i (a), but after filtration 6 to 8 per cent of glycerine 

 (sp. grav. 1.25) is added. This medium is especially used 

 for growing the tubercle bacillus when the soluble products 

 of the growth of the latter are required. 



2. Gelatine Media. These are simply the above 

 broths, with gelatine added as a solidifying body. 



2 (a). Peptone Gelatine. Take of meat extract say 

 1000 c.c., add 5 grammes sodium chloride, 10 grammes 

 peptone, and from 100 to 150 grammes gelatine (the "gold 

 label " gelatine of Coignet et Cie, Paris, is the best). The 

 gelatine is cut into small pieces, and added with the other 

 constituents to the extract ; they are then thoroughly 

 melted on a sand bath, or in the " Koch." The fluid 

 medium is then rendered slightly alkaline, as in i (a\ and 

 filtered through filter paper. As the medium must not be 

 allowed to solidify during the process, it must be kept 

 warm. This is effected by putting the flask and funnel 

 into a tall Koch's steriliser, in which case the funnel 

 must be supported on a tripod, as there is great danger 

 of the neck of the flask breaking if it has to support the 

 funnel and its contents. The filtration may also be carried 

 out in a hot-water funnel (Fig. 7). This consists of an 



