264 PRINCIPLES AND PRACTICE OF MILK HYGIENE 
less time, and is found to be equally satisfactory for 
plating, as explained below. 
“ Sterilize in an autoclav at 15 lbs. pressure (never 
above) for one-half hour after dry steam comes from the 
autoclav. As an alternative the medium may be steri- 
lized on three successive days in streaming steam, in 
which it must remain 20 minutes each day after the agar 
has completely melted. 
“Tf the flasks are to be kept for more than a couple 
of days, or are to be shipped, before using, cover the 
mouths of the flasks before sterilizing with paper fastened 
by a rubber band or by tinfoil to retard evaporation.” 
Dilutions— “For miscellaneous milk samples the 
character of which is not known three dilutions shall be 
made: 1-100, 1—1000, 1-10,000. Where the character 
of the milk is known the number of dilutions may be 
reduced. If the milk is pasteurized, certified, or 
known to be fresh and of high grade, the 10,000 
and 1000 dilutions may be omitted; if the milk is known 
to be old and of high bacterial content the 100 dilution 
may be omitted. In no case shall less than two plates 
be made of each sample. 
“ Any convenient method of making dilutions may 
be used, always using pipettes and sterile water blanks. 
The water used for dilutions may be placed in dilution 
bottles (99 cc. and 9 cc. are convenient sizes) and 
sterilized for one hour in an autoclav at 15 Ibs. pressure. 
These should be marked so that it can be determined that 
they have neither gained nor lost water during or subse- 
quent to sterilization. Or the water may be sterilized 
in bulk, if kept in a properly guarded container, and 
subsequently measured directly into dilution bottles with 
sterilized pipettes. 
