CULTURE MEDIA 47 



filtered and then rendered plus i per cent, alkaline. With the above solutions, 

 the medium is made as follows: 



Place 300 cc. of the 10 per cent, egg-white solution in a liter flask; 300 cc. 

 of the 10 per cent, egg-yolk solution in another flask, and 400 cc. of the meat 

 infusion, to which is added 15 Gm. of powdered agar-agar, in a third flask. 

 These are then sterilized in the autoclave at 15 pounds pressure for 15 minutes. 

 They are removed from the sterilizer and, while hot, i cc. of a i per cent, alco- 

 holic solution of gentian violet is added to the broth-agar. The contents of this 

 flask are now poured into that containing the egg white and then the egg yolk 

 is added. The whole is poured back and forth from this flask to another so as to 

 insure thorough mixing, and then it is tubed and slanted. The tubes are left 

 in slanted position for about 72 hours, at room temperature, until the contents 

 are well set, and the tubes sealed with corks or paraffin. 



REACTION OF CULTURE MEDIA 



The most common organisms which are both saprophytic and pathogenic 

 flourish on media either slightly acid, neutral or slightly alkaline in reaction. 

 Many of these grow most luxuriantly in a medium having a reaction indicated 

 as slightly alkaline when tested with litmus paper (turn red paper slightly blue, 

 do not turn blue paper red). For general purposes, media tested with litmus 

 paper and adjusted to give a slightly alkaline reaction, do well. 



Plain bouillon, agar and gelatin media, when made, are usually acid to litmus 

 paper. When it is necessary to add acid or alkali, solutions of hydrochloric acid 

 or sodium hydrate are used. 



Although this easy method of testing and adjusting the reaction of culture 

 media is ordinarily satisfactory, it has faults that make more accurate methods 

 preferable, at least for some purposes. The observations of different workers 

 are comparable only when the culture media they use is uniform in reaction. 

 This is especially true and important in culturing milk, water, sewage and food- 

 stuffs to determine the number of bacteria contained. For this reason the 

 American Public Health Association has recommended the following method 

 of testing and adjusting the reaction of culture media: 



Reagents freshly prepared % per cent, solution of phenolphthalein in 

 50 per cent, alcohol indicator. 



Normal, tenth normal and twentieth normal solutions of hydrochloric acid 

 and sodium hydrate. 



Place 5 cc. of culture medium in porcelain dish, add 45 cc. distilled water, 

 boil for 3 minutes, add i cc. of phenolphthalein (indicator) ; if acid, the contents 

 of the dish show no change of color; if alkaline, turn red. Usually the reac- 

 tion is acid; in this case tenth normal NaOH is added, drop by drop, from a 

 graduated burette until the neutral point is reached; the amount of NaOH 

 required to neutralize is recorded, and the experiment repeated, using twentieth 

 normal NaOH. 



Should the contents of the dish turn red when the phenolphthalein is added 

 it is alkaline and titration would be done with ^ and ? HC1. 



