34 AIDS TO BACTERIOLOGY 



hydrochloric acid, and well rinsed with water to remove 

 all traces of acid. The tubes are then allowed to drain 

 until nearly dry, when they are finally rinsed out with a 

 little strong alcohol, drained, and allowed to dry. Or the 

 tubes may be cleaned by boiling in water containing soap 

 powder for thirty minutes, followed by cleansing with 

 brush and thorough rinsing with water. They are then 

 plugged with sterilised cotton-wool, and sterilised for an 

 hour at 140 to 150 C. in the hot-air steriliser. Cotton- 

 wool is sterilised by pulling loosely apart and heating for 

 an hour at 145 C. in the hot-air steriliser. Though 

 sometimes desirable, it is not always necessary to sterilise 

 tubes and cotton-wool before filling with media. The 

 sterilisation after tubing is generally sufficient. 



Reaction of Media. A reaction slightly acid to phenol- 

 phthalein (equivalent to a faintly alkaline reaction to 

 litmus) is generally most suitable. 



Standardisation of Media. Variations in reaction of 

 media influence the character of the growths. For the 

 enumeration of organisms and for descriptive work 

 standard media are necessary. The signs + and indicate 

 acidity and alkalinity to phenolphthalein respectively. 

 The American Committee describe +1-5 per cent, reaction, 

 when to every 100 c.c. of medium neutral to phenolphtha- 

 lein 1 '5 c.c. of normal hydrochloric acid are added. English 

 workers, following Eyre, use an acidity of +1-0 per cent. 

 (+ 10 on Eyre's scale). 



The procedure is briefly as follows: An aliquot portion 

 of the nutrient medium is taken, say 20 c.c. ; this is diluted 

 with warm distilled water, boiled for a minute, a few 

 drops of a solution of phenolphthalein are added, and 

 decinormal solution of sodium hydrate is run in drop by 

 drop from a burette to the hot solution until a pink colora- 

 tion is obtained. The correct volume in c.c. of normal, or, 

 better, dekanormal, soda solution to be added to the bulk 

 is calculated and added; the reaction of the medium will 

 then be neutral to phenolphthalein, but strongly alkaline 

 to litmus. The alkalinity is too great for the optimum 

 growth of most organisms, and it is reduced by the addition 

 of normal hydrochloric acid to the extent of 1 c.c. per 100 

 c.c. of medium. The reaction is then said to be + 10 

 (Eyre's scale) or 4- 1*0 per cent. (American scale). 



Instead of first neutralising and then adding normal 





