MILK-BORNE INFECTIONS 451 



After inoculation tiie flasks are incubated at 37° C. When 

 the surface is covered with a heavy film the fluid is shaken care- 

 fully so as to cause the film to sink, leaving enough surface growth 

 to start a new generation. This process is repeated several times. 

 The length of time required for sufficient growth is from four to 

 twelve weeks. When freshly isolated strains are used the incuba- 

 tion period is longer than when strains have been recultivated for 

 some time and have become habituated to the medium. Viru- 

 lence of the organism is gradually lost, but the products are such 

 as to produce an efficient tuberculin. After the incubation period 

 the flasks are placed in a hot-air oven until the fluid boils. This 

 destroys all living bacilli. The cotton plugs are removed, the 

 fluid filtered, and evaporated over a water-bath. Koch evapo- 

 rated his tuberculin to one-tenth the original volume. When the 

 desired volume has been attained, 0.5 per cent, carbolic acid is 

 added as a preservative. For use on human beings tuberculin is 

 finally filtered through a Berkefeld filter for complete sterilization. 

 The finished product is kept in sealed bottles in the dark. 



Standardization of tuberculin is not very satisfactory, but 

 several methods are in use. The amount of tuberculin necessary 

 to produce a rise of 4° to 5° C. in a guinea-pig of 1 pound weight 

 forms one standard; by another the amount of tuberculin neces- 

 sary to kill a tuberculous guinea-pig is determined. Sometimes 

 the amount of acid produced by the tubercle bacillus serves as a 

 guide. It is assumed that the amount of acid corresponds to the 

 growth that has taken place. 



It takes considerable experience to make a test with tubercu- 

 lin for tuberculosis in cattle. The normal temperature of cattle 

 is not always the same, varying from 1 to 3 degrees, according 

 to such circumstances as warm weather, the use of cold drinking- 

 water, or to disturbances in health. The temperature in dif- 

 ferent individuals is not quite uniform. The average for cows 

 is 101° to 102° F., probably nearer to 102° F. in the majority of 

 cases. The best season for the test is when the weather is cool 

 and the animals are kept most of the time in stables. The tem- 

 perature of the animal to be tested is taken during the day every 

 two or three hours and the tuberculin injected in the evening of 

 the same day. Commencing early the next morning the tem- 

 perature is taken again at regular intervals. The temperatures of 

 the previous day and of the day following injection are then 

 plotted on a chart and the curves compared. A sample of such 

 a chart is shown in Fig. 190. 



If the maximum temperature after injection is 2° F. or more 

 above the normal temperature, tuberculosis is indicated. A rise 

 of 1.5 degrees is suspicious, and the animal should be retested 



