NUMBER AND SOURCE OF BACTERIA IN MILK 257 



Ex. 125. Determine the number of bacteria in a sample of 

 milk. Since the number of bacteria is usually very large in an 

 ordinary sample of milk, it is necessary to dilute the latter in 

 order to avoid too much crowding of the colonies on the plates 

 which are inoculated by it. 



For the purpose of dilution sterile water must be used. 



It is usual to dilute the milk 10, 100, 500, or 1000 times. 



Proceed as follows : 



From a burette run into a series of test-tubes 9 c.c. of water, 

 and mark the level of the water in each tube with a file ; add a 

 few drops more, so that the water stands a little above the mark. 

 Plug with cotton-wool, and sterilize by heating on three suc- 

 cessive days for thirty minutes. (It will be found useful to 

 have a series of small conical flasks marked at the 49 c.c. level, 

 and a similar lot marked at the 99 c.c. level.) 



Now take the sample of milk to be examined and shake it 

 vigorously to obtain uniform distribution of its contents. 



Remove with a sterile i c.c. pipette i c.c. of milk, and add it 

 to the test-tube containing 9 c.c. of water. The milk is now 

 diluted 10 times (i in 10), and i c.c. of the mixture of milk and 

 water contains -^ of a c.c. of milk. 



Gently shake the mixture for a short time, and then remove 

 with another sterile pipette i c.c. from this first dilution and 

 add it to another tube containing 9 c.c. of sterile water. The 

 second dilution now obtained is i in 100, and T c.c. of it 

 contains JJ^ c.c. of the original milk. Shake and transfer i c.c. 

 of this to 9 c.c. of sterile water, the dilution in the third tube is 

 i in 1000, i c.c. containing T oVo c - c - of the milk. 



If a dilution of 500 times (i in 500) is required, add i c.c. 

 to 9 c.c. of water, and then i c.c. of this to a flask containing 

 49 c.c. of water. 



Having prepared a suitable dilution, say i in 1000, add .5 c.c. 

 of it to a tube of melted lactose gelatine from a sterilized 

 graduated pipette, and, after gentle mixing, pour the inoculated 

 medium into a Petri dish and allow it to solidify. 



