No. I.] SYMPATHETIC GANGLIA OF VERTEBRATES. 31 



The methylene blue method, as used by me in this research, 

 may be briefly described as follows: A i^, 2^, or 4^ solution 

 of methylene blue (Grubler's rectificiert nach Ehrlich), made in 

 a normal salt solution, was injected into a vein, easily accessible. 

 In frogs, the large lateral cutaneous vein ; in Reptilia, the jugu- 

 lar ; in birds, the humeral (Owen); in Mammalia, the jugular 

 or femoral, usually the former, were used. As to the percent- 

 age of the solution used, no definite statement can be made ; 

 the above solutions were used with equally good results in some 

 experiments, and equally unsatisfactory results in others. In a 

 general way it may be said that the stronger solutions stained 

 more readily the cell body and branches of the sympathetic 

 neurons, while the weaker solutions brought to view more 

 clearly the pericellular plexuses ; these statements are, how- 

 ever, open to many exceptions. 



The quantity injected varied with the size of the animal 

 used ; from 2 to 4 c.cm. for a frog, to 60 to 80 c.cm. for a 

 dog. The solution was allowed to flow into the circulation 

 until the animal became blue, or until the heart's action was 

 stopped. 



Forty-five minutes to one hour after the injection, the gan- 

 glia or tissues to be examined were exposed ; the larger ganglia, 

 with the afferent and efferent nerves, were freed from the sur- 

 rounding tissues, but not removed until they assumed a blue 

 color, when they were excised and placed on a slide, and, if 

 the staining seemed satisfactory, were placed in the fixative. 

 Smaller ganglia were at once removed to a slide moistened with 

 normal salt, and examined from time to time, until the stain 

 was developed, when they were also placed in the fixative. The 

 fixative used by me had the following composition : 



Ammonium molybdate i grm. 



Aqua dest 10 c.cm. 



Hydrochloric acid i gtt. 



The molybdate is ground in a mortar and the water added ; 

 the solution is then removed to a flask and heated until per- 

 fectly clear ; the hydrochloric acid is then added. The solution 

 so made is placed in small glass jars, and these are surrounded 



