288 PRACTICAL DAIRY BACTERIOLOGY 



with the above stain, and allow it to stain for five minutes, 

 warming slightly. Wash off the superfluous stain, and place in 

 the acid solution, until all traces of red have been removed, 

 which will require, usually, from about three to five minutes. 

 Wash off the acid, dry and flood the slide for about half a 

 minute with methylene blue solution. Wash again, dry and 

 examine with a compound microscope, without a cover-glass. 

 The tubercle bacilli will be found, by this method, to stain red, 

 whereas other bacteria will be stained blue. It requires a little 

 experience to be successful with this method of staining, but 

 it is fairly easy after a little practice. 



*No. 20. Qualitative Bacteriological Analysis. This is very 

 difficult and can be successfully undertaken only after consider- 

 able experience. A qualitative analysis of milk must begin with 

 an estimation of the number of bacteria to be expected, in order 

 that a proper dilution may be selected to give 200 to 1,000 col- 

 onies per plate. If the age and temperature of the milk are 

 known, an approximate guess may be made. If the milk is less 

 than 12 hours old and is kept moderately cool, a dilution of 100 

 and 1,000 may be chosen. If from 12 to 24 hours old, 1,000 to 

 10,000 dilution will be better. If the milk is kept as warm as 

 2OC higher dilutions are needed, sometimes as high as 1,000,000 

 in warm milk 24 to 36 hours old. If it is kept cool, 50^ but if 

 48 hours old, it will be likely to require 10,000 to 30,000. These 

 numbers are, unfortunately, not very definite. 



Secure some milk not more than 6 hours old. Dilute 1,000 

 times and make several litmus gelatin plates exactly as in ex- 

 periment ii. 



It is always best to use two or three different dilutions in 

 order, if possible, to insure proper results in one set, at least. 

 At the end of two days examine the plates to see whether rap- 

 idly liquefying colonies are liable to spoil the plates. If lique- 

 fiers are abundant the plates must be studied at once; but if not, 

 it is best to leave them for from 4 to 6 days before study. 



