210 MICROSCOPIC AND CULTURAL STUDY OF BACTERIA 
are conveniently prepared from a layer of cotton batting about 2 
inches square (for a test-tube of ordinary diameter, about 15 mm.), 
which is laid squarely over the orifice. The center of the square is 
gently pushed down into the neck of the tube for a distance of about 
f to 1 inch ; sufficient cotton protrudes from the tube to be conveniently 
grasped by the fingers and removed. It is frequently advisable to 
cover the cotton plugs after they are in place with two or three layers 
of filter paper, which prevents an accumulation of dust on the cotton. 
Wide-mouthed containers are sealed with several layers of unglazed 
paper fastened in place wuth a piece of twine. Flasks are frequently 
not plugged with cotton; the neck is simply covered by an inverted 
beaker of appropriate size. 
Glassware should always be sterilized before media is placed in it; 
this is readily accomplished by dry heat. A hot-air sterilizer is used, 
in which a temperature of 180° C. is maintained for one hour. A 
higher temperature must be avoided, to prevent charring of cotton 
plugs. The heat must be increased gradually and diminished gradu- 
ally, to prevent cracking of the glass. By this process not only is 
the utensil rendered sterile, the plugs of cotton retain their shape when 
wuthdraw^n, as well. 
A majority of the bacteria pathogenic for man and many parasitic 
and saprophytic forms as well, require relatively complex organic 
compounds containing carbon, hydrogen, nitrogen and oxygen, 
together with other elements for their nutrition. These foodstufi^s 
provide both the structural and fuel requirements of the organism, 
as explained in the chapter on Bacterial Metabolism. Experience has 
shown that a medium containing meat infusion, peptone, and salt is a 
satisfactory one for many bacteria. This medium may be enriched 
by the addition of various ingredients to meet the requirements of the 
more fastidious organisms. 
Preparation of Meat Infusion.— Meat infusion is prepared from finely 
comminuted lean meat^ freed from fat. Five hundred grams of meat 
are intimately mixed with 1000 cc. water and allowed to infuse over 
night in the refrigerator. It is then strained through several layers of 
cheese-cloth, the volume recorded, then heated to boiling. The 
coagulum which forms is removed by filtration through filter paper 
and the clear amber-colored fluid, after restoring the loss due to 
evaporation, is run into flasks and sterilized in an autoclave at 15- 
pounds' pressure for fifteen minutes. This plain meat infusion 
contains but little protein; it is relatively rich, however, in soluble 
meat extractives, soluble salts and muscle-sugar— glucose. It is not 
suitable in itself as a complete nutritive medium for most bacteria, 
but it forms the basis of many of the commonly used nutritive media. 
Meat extract (Liebig's or other kinds) is frequently substituted for 
meat infusion. Three grams of meat extract are dissolved in a small 
' Beef hearts make a very satisfactory meat infusion and their cost is much less than 
the better cuts of meat. 
