36 METHODS OF CULTIVATION OF BACTERIA 



determined, and the optimum concentration for bacterial growth is 

 unknown, though probably it . approximates that of the blood serum. 

 A still more serious difficulty is that no simple means is available for 

 estimating the hydrogen-ion concentration of many media, e.g., agar. 

 It is nevertheless true that although the methods in use are largely 

 empirical the products resulting are quite satisfactory for all but the 

 more delicate bacteria dealt with. It is therefore probable that many 

 i organisms will tolerate a reaction which may extend from a point some 

 way on the alkaline side of the optimum to a point some way on the 

 acid side. Precise statements such as are frequently made- regarding the 

 adjustment of a medium to a certain reaction — say, on Eyre's scale—do 

 not represent anything else than an expression of the fact that by the 

 technique practised a suitable medium has been produced. It has not 

 been usually recognised, for instance, that hydrogen-ion concentration is 

 a function of the temperature at which the reaction has been adjusted. 

 Thus, a reaction of, say, +10 adjusted at the boiling-point is no longer 

 a +10 reaction at the incubation temperature at which the medium is 

 actually used. 



1 (b). Glucose Broth. — To the other constituents of 1 (a) 

 there is added 1 or 2 per cent, of glucose. 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 (c). Glycerin Broth. — The initial steps are the same as in 

 1 (a), but after filtration 6 to 8 per cent, of glycerin (sp. grav. 

 1 - 25) is added. This medium is especially used for growing the 

 tubercle bacillus when the products of the growth of the latter 

 are required. 



2. Gelatin Media. — These are simply the above broths, with 

 gelatin added as a solidifying body. 



2 (a). Peptone Gelatin : — 



Meat extract .... 1000 c.c. 

 Sodium chloride .... 5 grms. 



Peptone albumin . . . . 10 „ 



Gelatin .... 100-150 „ 



(The " gold label " gelatin of Coignet et Cie, Paris, is the best.) 

 The gelatin 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 1 (a), and filtered through filter paper. 

 As the medium must not be allowed to solidify during the pro- 

 cess, 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 or diaphragm, 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 



