256 Value of Antiseptics 



6. The Culture-media used for the primary culture, and for the 

 subcultures, made after exposure of the micro-organism to the dis- 

 infectant, is nutrient bouillon made with Leibig's beef extract in 

 the usual manner and given a reaction of exactly + 1.5. Anderson 

 and McClintic achieve this by so carrying out the titrating of the 

 medium that a distinctly perceptible pink color marks the point at 

 which the addition of the alkali stops (see directions for titrating 

 culture-media). 



7. The Tubes for the culture and subcultures are ordinary culture 

 tubes, containing 5 cc. of the nutrient bouillon mentioned above. 

 They are filled, plugged and sterilized in the usual manner. 



The tubes for "seeding," i.e., exposing the bacteria to the ger- 

 micide, are more convenient when shorter. At the time of transfer, 

 the platinum loop is to be introduced into the tube as it stands in 

 the water-bath and as this is not easy with tubes of standard length, 

 Anderson and McClintic recommend tubes i inch in diameter and 

 3 inches long. These are plugged and sterilized by dry heat, 

 or as recommended by the authors quoted, are sterilized mouth 

 down, without plugs in a paper-lined wire basket. 



8. The Dilution of the Phenol and Test Solutions. This is done in 

 standardized graduates with standardized pipets, according to 

 the requirements of the particular case. Anderson and McClintic 

 give tables that are useful for making the dilutions, though with the 

 aid of a little arithmetic it is easy to calculate the proportions of 

 the 5 per cent, solutions already prepared, and sterile distilled water 

 necessary to make the test solutions required. As it is certain that 

 some of the dilutions will be below germicidal strength, and as 

 " weeds" may be more difficult to kill than the test organism (B. 

 typhosus) it is important to see that the distilled water used for 

 dilution is sterile, and that the cylinders and bottles or pipets 

 used for making the dilutions are all sterile and that the dilutions 

 themselves are made with aseptic precautions. 



Under the standard conditions recommended, the phenol solu- 

 tion that destroys all of the B. typhosus introduced, in 2% minutes 

 is i : 80, but it is always wise to make additional dilutions to control 

 the strength, as shown in the table below. When the strength 

 of the disinfectant or germicide to be tested is entirely unknown, 

 it is well to begin by making a number of tests with widely sepa- 

 rated dilutions, by one of the "rough and ready" methods, so as 

 to arrive at an approximate strength, before commencing the more 

 difficult technic required for the determination of the phenol co- 

 efficient, which should be looked upon as the final test for exact 

 comparison. 



9. Racks for Holding the Tubes are indispensable. The "seeding 

 tubes," that is, the tubes in which the actual exposure of the culture 

 to the germicidal solutions is to take place, have already been pro- 

 vided for in the construction of the water-bath. 



