METHODS FOR MICROSCOPIC STUDY OF BACTERIA 227 
indol by the splitting off of alanin from the tryptophan molecule: 
(see page 72). 
/\ /\ 
\ CH2.CHNH2.COOH / \ 
I [ =1 11+ Alanin. The alinin is 
I , I III decomposed by the 
/\/ \ /\/ bacteria. 
\/ NH \/ NH 
Tryptophan. Indol. 
Test for Indol —ImM in mhieral acid solution (IICl, or TI2SO4) 
gives a red color with nitrites. 
r 
r 
HNO2 
NH \/ NH 
Nitroso indol. 
Nitroso skatol and nitroso compounds of derivatives of indol, do 
not give the red color. 
Samples of peptone occasionally do not contain tryptophan, con- 
sequently each lot of peptone should be tested. When an especially 
favorable sample is found it should be reserved for this purpose. 
Plain neutral sugar-free broth is a better medium than Dunham's 
solution for the indol test, and it should be employed for this purpose 
whenever possible. 
Nitrate Broth.— Add 10 gm. of peptone of 1 liter of water and dis- 
solve by boiling. Then add 0.2 gm. chemically pure potassium nitrate 
—free from nitrites — and filter. Sterilize in the autoclave. The 
reaction does not require adjustment. 
Nutrient Gelatin Media.— Ten grams of peptone and 5 gm. of XaCl 
are added to 1 liter of sugar-free meat infusion^ and dissolved by boiling. 
When the ingredients are in solution, 100 gm.- of the best quality 
gelatin are added, a few leaves at a time, and stirred until dissolved. 
The reaction is then adjusted to the desired degree and verified after 
an additional five minutes' heating. The medium is cooled to 50° C, 
and clarified with eggs, using 2 eggs for each liter. Filter through a 
double layer of absorbent cotton in a large glass funnel until clear, and 
sterilize. When sterilization is accomplished, cool quickly and store in 
the ice-box. 
Nntrient Agar. — (a) Dissolve 12 gm. of powdered or shredded agar 
in 1 liter of meat infusion by the aid of heat, and add 5 gm. XaCl and 
10 gm. of peptone. Maintain a boiling temperature for at least thirty 
minutes, or until the ingredients are completely dissolved, restore the 
loss by evaporation, adjust the reaction, and filter through a double 
' Meat extract may be used in place of meat infusion, but the medium is not as satis- 
factory for pathogenic bacteria. 
2 Use 120 gm. gelatin during warm weather. 
