218 MICROSCOPIC AND CULTURAL STUDY OF BACTERIA 
of ordinary bacteria. The addition of 10 cc. of normal acid to a liter 
of medium of this alkalinity, containing the usual amount of phosphate, 
would reduce the pH value to approximately pii 7.(). This is very 
nearly the pH value of the blood and therefore well suited for the 
growth of pathogenic bacteria. 
If, therefore, the titration of the medium is such as to require 30 
cc. of Y NaOH to bring the reaction to the turning-point of phenol- 
phthalein (pH 8.3), the addition of 20 cc. of y XaOH should result 
in a pH value of approximately pn 7.6, the point desired. A second 
titration, after the alkali is added, is desirable. If necessary, a second 
readjustment of the reaction may be made on the basis of the rede- 
termination. 
The Clarification of Media.— It is desirable, in the preparation of 
many cultm-e media, including broth, agar and gelatin, to remov^e all 
insoluble substances. This is accomplished by filtration methods, with 
or without preliminary treatment, to flocculate the substances in sus- 
pension. The addition of non-heat-coagulable proteins to broth, as for 
example, gelatin, frequently requires clarification with a coagulable 
protein, as egg-albumen, to remove the finely divided suspended 
matter. 
Clarifying with Eggs.— For each liter of medium to be clarified, 2 
eggs, thoroughly whipped in a small amount of water are added. The 
temperature of the medium should not exceed 50° C. The eggs are 
thoroughly stirred in and the entire mixture is slowly heated at 100° 
C, either in a double boiler or in the Arnold sterilizer. A firm coagu- 
lum forms during the heating process, which enmeshes the suspended 
particles it is desired to remove. The medium should never be 
disturbed during the coagulating process. The clear underlying 
medium is drawn oft' and filtered through cotton. 
Filtration Through Cotton.— A large glass funnel is lined with a double 
layer of absorbent cotton; the layers are placed at right angles, thus 
crossing the fibers of cotton at right angles. The cotton is moistened 
with a small amount of hot water if agar or gelatin is to be filtered. 
The medium is then carefully poured into the funnel, care being taken 
that the cotton is not displaced by the force of the inflowing fluid. 
The first portion of filtrate may not be clear and it is somewhat diluted 
with the water originally used to wet the cotton— hence it should be 
returned and refiltered. Agar and gelatin filter slowdy, which may 
lead to congelation, therefore the top of the funnel should be covered 
to prevent undue loss of heat. Funnels surrounded by a hot-water 
jacket are sometimes used in the filtration of these media. Media 
that are fluid when cold may be often advantageously clarified by 
filtration through a good grade of heavy filter paper, with or without 
a preliminary clarification with eggs, as occasion demands. 
The Distribution of Media.— The clarified media are either stored in 
flasks or transferred to smaller containers for immediate use. Then 
thev are sterilized. Most media are used in test-tubes. Test-tubes 
