PREPARATION OF MEDIA 45 



properties and some contain intact yeast cells which may be misleading 

 in microscopic preparations. 



Semisolid agar. Some organisms, especially microaerophiles, are 

 cultivated more successfully in semisolid media in which agar, in concen- 

 trations varying from 0.2 per cent to 0.5 per cent according to the purpose 

 for which the medium is intended, is added to a suitable base medium. 

 Such a medium may be useful also for determination of motility and for 

 fermentation reactions. To aid in reducing the degree of oxidation of 

 the medium the lower concentration of agar is sufficient; for determina- 

 tion of motility, concentrations approaching 0.5 per cent are necessary in 

 order to stiffen the medium sufficiently to show distinctly the hazy zone 

 characteristic of motile bacteria (Tittsler and Sandholzer, 1936). Since 

 the concentration for this purpose is critical and the percentage to be 

 used will vary with the purity of the agar, the exact concentration to be 

 used must be tested for each batch of agar and each lot of medium checked 

 with a known motile culture. 



Silica gel. It is sometimes desired to cultivate bacteria on inorganic 

 gels to avoid unknown and possibly undesirable chemical contaminants 

 in agar, to avoid liquefaction of agar by certain types, and to eliminate 

 agar in the cultivation of certain autotrophic bacteria. Sterges (1942a 

 and b) published full details (which cannot be condensed here) of prepara- 

 tion of such media based on the reaction between sodium silicate and 

 hydrochloric acid. A simplified technic recommended by Ingelman and 

 Laurell (1947) follows: Mix 1 vol of ortho-silicic acid tetraethyl ester, 

 Si(OC2H5)4, with 1 vol of ethanol; add 6 vol of boiled water a little at a 

 time with stirring. Remove turbidity by centrifuging, and dispense in 

 tubes or plates. Sterilize at 120°C for 30-40 min in autoclave, during 

 which time gel formation occurs. Cool slowly, and remove ethanol by 

 flooding with sterile water. Remove water, and replace with suitable 

 nutrient solution. Reautoclave if necessary for sterility. Temple (1949) 

 recommends a commercially available, highly purified colloidal silica 

 preparation such as Ludox (duPont) ; for use a 30 per cent aqueous solu- 

 tion is diluted to 10 per cent, nutrients are added, pH adjusted, it is dis- 

 pensed in petri dishes and autoclaved. 



Storage media include those media in which bacteria are stored in 

 "stock culture" condition for indefinite periods to provide a source of 

 viable cultures as needed. The medium to be used will vary with the 

 species to be maintained. A critical factor to be considered is whether or 

 not the presence of a fermentable carbohydrate reduces viability; fre- 

 quently 2 per cent CaCOa (sterilized separately by dry heat) is added to a 

 medium to neutralize the acids produced by an organism which will not 

 grow well in the absence of a fermentable carbohydrate. After incuba- 

 tion in the chosen medium at the optimum temperature for a period 



