304 



DISINFECTANTS AND ANTISEPTICS 



tion with deci-normal bromine water as described under "Phenol" in the United States 

 Pharmacopoeia (loth rev.), or with sodium bromide and bromate solution as described by- 

 Francis Sutton.' Preserve it in 200-cc. amber-colored tightly stoppered bottles, protected 

 from the light. 



Proportion of culture to disinfectant. — Five-tenths cc. of unfiltered culture is added to 

 S cc. of disinfectant. 



Inoculation loops. — A 4-mm. loop of platinum wire, United States 24 standard gauge, 

 i| inches long, is set into any suitable holder, such as aluminum or glass rod about 0.5 cm. in 

 diameter. 



Incubation. — Subcultures are incubated for 48 hours at 37 °C. 



Dilutions. — Any series of dilutions which the operator considers advisable may be used, 

 but these dilutions must be made accurately and, within the limitation of the test, must show 

 the maximum dilution capable of killing B. typhosus in 10 minutes. The method of making 

 the dilutions also is left to the operator, as it is presumed that he has sufficient training to do 

 this accurately. 



Seeding tiibcs. — Lipped test tubes 5XI inches, plugged with cotton, as used in the 

 Rideal-Walker method, or test tubes of even larger size, according to the preference of the 

 operator, may be used for medication. 



Subculture tube racks. — The subculture tube racks specified in Hygienic Laboratory Bull. 

 82 are convenient. 



Method of conducting the test.- — Any number of dilutions up to ten may be used. Of these 

 dilutions two must be reserved for the phenol control. Dilutions of 1-90 and i-ioo phenol 

 are arbitrarily chosen for this purpose. These are accurately made from the stock 5 per 

 cent solution of phenol. 



At intervals of 30 seconds, 0.5 cc. of culture is run into 5 cc. of each dilution from a 5-cc. 

 pipette. Care must be taken not to contaminate the sides of the tubes with the culture and 

 be sure that no cotton threads adhere to the open ends of the seeding tubes. After inoculating, 

 thoroughly sterilize in the flame the ends of the tubes where the pipette has touched. 



At intervals of 5, 10, and 15 minutes transfer from each dilution to the specified broth. 

 This will allow 30 seconds between transfers if ten dilutions are used. Incubate the tubes 

 48 hours at 37° C. and read results. If there is any doubt that the growth in the subcultures 

 is B. typhosus, this can be confirmed by agglutination tests. 



Calculation of coefficient. — Divide the greatest dilution of the disinfectant capable of 

 killing B. typhosus in 10 minutes but not in 5 minutes by the phenol dilution which should 

 do this and divide these figures one into another. In order not to convey a false idea of the 

 accuracy of the method the coefficient is calculated to the nearest o.i point if under i.o, to 

 the nearest 0.2 point if between i and 5.0, to the nearest 0.5 point if between 5 and 10, and 

 to the nearest i.o point if between 10 and 20. For example, if results are read as follows: 



DISINFECTANT 



Dilution 



1-300. 

 1-350 

 1-400 

 1-450 



5 Min. 



10 Min. 



15 Min. 



+ 

 + 



Sutton, Francis: A Systematic Ildndbook of Volumetric Analysis (nth ed.), pp. 404-5. 1924. 



