WATER 33 



sterilized in the hot-air sterilizer after thorough cleaning and 

 plugging of the upper ends with cotton wool. 



In practice proceed as follows : Melt agar and gelatine 

 tubes, and cool down to 42 or 43 C. for the agar, and 25 

 to 30 C. for the gelatine. Mix the sample thoroughly. Add 

 O'2 and 0*5 c.c. of the sample respectively to two gelatine 

 tubes, and O'2 and ro c.c. to agar media tubes. Distribute the 

 water uniformly through the medium by rotation between the 

 fingers. Pour out, after flaming the cotton-wool plugs, into 

 sterilized Petri dishes, only raising the upper dish just suffi- 

 ciently to admit the top of the test-tube. Solidify as soon as 

 possible either over ice or by using a plate-cooling apparatus 



(Fig- 5). 



Fig. 5. Savage's Plate-cooling Apparatus. 



The inlet pipe is connected by indiarubber tubing to a water tap and the outlet 

 pipe to a sink. Plates of media placed on the thick plate-glass top are rapidly 

 solidified. 



Incubate the gelatine plates at 20 to 22 C. and the agar 

 at 37 C. Count the agar plates the next day, and after forty 

 to forty-eight hours. Count the gelatine plates daily, the final 

 count being at the end of seventy-two hours. 



To count the colonies it is best to count against a dark back- 

 ground, dividing up the area of the plate, to facilitate counting, 

 by lines on the back made with a paraffin pencil. 



All the colonies on the plate should be counted, but if they 

 are very numerous, and an approximate estimation only is 

 possible, then, but only then, some mechanical aid such as 

 Pakes' disc may be used, a few segments counted, and the 

 total number deduced. 



In stating results, the details of temperature, time of incu- 

 bation, and reaction of the medium should always be given 

 and recorded with the analysis. 



s. w. 3 



