LABORATORY WORK 337 
No. 4. Determination of the Bacteria in Milk.—Milk commonly contains so 
many bacteria that it must be diluted before the bacteria are determined. The 
amount of dilution needed will vary widely with the age of the milk. For most 
market milk a dilution of 1,000 will serve. Proceed as follows: Into a 99 c.c. water 
blank place 1 c.c. of the milk to be tested. Shake vigorously so as to distribute the 
bacteria uniformly. With a second pipette transfer 1 c.c. of this mixture to a 9 c.c. 
water blank and again thoroughly mix. With a third pipette place 1 c.c. of thisin a 
Petri dish and then pour upon it the contents of anagar tube. Agitate as in the last 
experiment, allow to harden and incubate in the oven for twenty-four hours. Usea 
counting plate in counting the colonies. lf they are too numerous to count directly, 
count the number in one of the areas in the counting plate and multiply by the 
number of such areas covered by the whole plate. This will give the number of 
colonies on the plate, and this multiplied by 1,000, will give the number in the original 
milk. ‘There may be anywhere from a few thousand to several millions. It fre- 
quently happens that the dilution of 1,000 is insufficient, giving too many colonies 
on a plate. Higher dilutions are needed in such instances, but whether to make a 
higher dilution can only be determined by a knowledge of the age and temperature 
of the milk and by experience. Plates are most satisfactory when they contain 
between 30 and 300 colonies each. 
From these and from the plates of the following two experiments isolate and 
purify cultures as directed in No. 7. 
No. 5. Bacteria in the Air.—Pour the contents of several tubes of agar into 
sterile Petri dishes, replace the cover and allow to harden. Remove the cover 
from. one of them and allow it to remain open in the laboratory for two minutes. 
Then replace the cover and place in the incubating oven. Expose a second in the 
same way ina barn; a third out of doors; a fourth in a barn after hay has been thrown 
down in front of cattle. Other Petri dishes may be exposed in other localities. 
After twenty-four hours’ incubation count the number of colonies and compare the 
relative number of bacteria in the air at the different places. 
No. 6. Bacteria on the Fingers.—Pour a Petri dish as above directed and allow 
to harden. Remove the cover of one and touch it gently with the fingers in several 
places. The hands may then be thoroughly washed, and a second Petri dish treated 
in the same way. Incubate for twenty-four hours and see if bacterial colonies have 
grown where the fingers touched the agar. 
No. 7. Isolation and Purification of Bacteria.—Any of the plates above pre- 
pared will show after proper incubation a number of colonies. Comparing the 
different colonics noticeable differences will be seen between them. These differ- 
ences in the colonies commonly indicate different kinds of bacteria, for the same 
kind of bacteria produce, commonly, the same kinds of colonies. 
Isolation.—Sterilize a platinum needle (Fig. 61), in a flame until red-hot and after 
allowing it a few seconds to cool, dip the tip of it into one of the colonies. Transfer 
the bacteria adherent to the needle to one of the agar slant tubes by removing the 
plug and drawing the tip of the wire over the surface of the agar. Sterilize the needle 
again before laying it down. Label the tube with a gum label telling its source. 
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