304 R. R. BENSLEY 



b. The janus green method. For this method the same tech- 

 nique is followed as in the preceding neutral red method. A word 

 is necessary here, however, with regard to the dye. It is neces- 

 sary for this purpose to secure the janus green recommended by 

 Michaelis ('00) for staining intra vitam certain cell granulations. 

 This, Michaelis says, is 'diethylsafraninazodimethylanilin.' The 

 corresponding dimethyl dye is ineffective both for the purpose 

 for which Michaelis recommended janus green and for staining 

 the islets of Langerhans. In this country the correct compound 

 may be obtained from L, A. Metz and Company, New York. 

 Several samples of janus green obtained from agents and bearing 

 G. Griibler's label were worthless for this purpose. The janus 

 green is injected in the form of a 1 in 15,000 solution in isotonic 

 salt solution, or, for dogs and cats, in Ringer's solution. When 

 the whole pancreas has taken on a deep blue green color the in- 

 jection is complete. The pancreas is then covered up with the 

 intestines, etc., so as to exclude the air. Reduction of the dye 

 then proceeds rapidly and at the end of fifteen or twenty minutes 

 if the pancreas be examined it will be found to have a distinct 

 red color with a tinge of blue. This reduction must be checked 

 at the proper moment, either by exposure of the pancreas to the 

 air, or by the injection into the duct of either ammonium molyb- 

 date solution or a solution of potassiuni iodide. But, for fresh 

 study, it is simply necessary to divide the pancreas into sufficiently 

 small fragments so that oxygen from the air may have free access 

 to all parts. Such a preparation shows the islets deeply blue on a 

 red background. This method is not so suitable for total counts 

 because, as Michaelis has pointed out, the reduction of the dye is 

 accomplished by the splitting off of the dimethylanilin group 

 leaving a safranin which cannot be reoxidized to the original form, 

 as in the case of the neutral red and methylene blue leucobases. 



For the cat and dog where the size of the pancreas is so great 

 as to preclude the possibility of a total count within the time limits 

 permitted by the rate of reduction of the dye, the janus green 

 method is preferable under proper precautions to the neutral red 

 method for establishing ratios between the numbers of islets and 

 the weight of the pancreas containing them, with a view to esti- 



