CULTIVATION OF BACTERIA. 137 while impure peptone gives a red or reddish-yellow pre- cipitate. Both the peptone and copper solution should be in a dilute form to make successful tests. The addition of 4 c.cm. of the following solution Rosalie acid, 0.5, 80 per cent, alcohol, 100. makes it become an excellent reagent for the detection of acids and alkalies. The solution is pale rose in color. If the bacterium produces acids, the color fades; if alka- lies, it intensifies. As the color of rosalic acid is destroyed by glucose, it cannot be used in culture-media contain- ing it. Theobald Smith calls attention to the fact that Dun- ham's solution is unsuited to the growth of many bac- teria, some failing altogether to grow in it, and recom- mends that, instead, bouillon free of dextrose shall be used. All bacteria grow well in it, and the indol-reaction is pronounced in sixteen-hour-old cultures. His method of preparation1 is as follows: beef-infusion, prepared either by extracting in the cold or at 60° C, is inoculated in the evening with a rich fluid culture of some acid-pro- ducing bacterium (Bacillus coli), and placed in the ther- mostat. Early next morning the infusion, covered with a thin layer of froth, is boiled, filtered, peptone and salt added and the neutralization and sterilization carried on as usual, To test for the presence of indol, the bacterium is planted in the culture-medium, allowed to grow for upward of twelve hours, and then subjected to the com- bined action of a nitrite and chemically pure sulphuric acid. In making the test, Smith adds to each tube i c. cm. of a o.oi per cent, solution of KNO2, freshly prepared, and 10 drops of chemically pure H2SO4. The presence of indol is characterized by the production of a red color. 1 Journal of Exp. Medicine, vi., Sept, 5, 1897, p. 546.