122 ALAN C. SUTTON 



ity of that particular method for the demonstration of the 

 structure in question. 



I aimed to follow Wilson's directions for the methylene blue 

 method as closely as possible with the few modifications I found 

 it advisable to make in using it with embryos. Instead of the 

 previous erratic results with the gold chloride technique, I soon 

 found that the nerve endings could be demonstrated in practically 

 every case. The method briefly is this : Keep on hand a stock of 

 0.5 per cent methylene blue (Ehrlich) in distilled water. From 

 this a fresh solution is made each day for injection b}^ diluting 

 one part of the stock solution with nine parts of physiological 

 salt solution. The injection flask filled wth the fluid is kept in 

 the thermostat at 40°C. It was found best to keep it at this 

 higher temperature since it is bound to drop a few degrees while 

 being used. "W^hile a variation of a few degrees does not seem to 

 matter, it is best to aim to have the solution reach the embryo 

 at about 37°C. In order to keep ttie embryos warm it is well to 

 have the room verj^ warm, and besides, either keep the uterus in 

 warm water or wrapped in warm towels. While injecting the 

 fluid it is advisable to have the flask arranged so that it can be 

 readily raised or lowered. A rope and pulley is as good a way as 

 any. In this way the pressure can be regulated very easily. 

 When everything is ready, put a glass cannula in the umbilical 

 vein, after previously laying a loose ligature — ready to be tied 

 quickly — around the umbilical arteries. Then connect the cannula 

 to the flask with glass tubing, using rubber tubing only when ab- 

 solutely necessary for connections, and then taking pains to bring 

 the glass ends as close together as possible. Now cut across the 

 umbilical arteries and start the methylene blue at a low pressure 

 (18 inches) — to wash out all the blood. When the fluid coming 

 from the arteries is bright blue, the ligatures already in place are 

 tightened up to stop the flow from the vessels. Continue to in- 

 ject — raising the pressure gradually until the tissues are so dis- 

 tended that the thin skin of the embryo is just ready to break. 

 With the small sizes it is necessary to inject them while the 

 embryo still lies in the amniotic fluid, with the amnion intact, 

 for if the embr3^o is handled the skin is bound to break in places, 



