41 6 Journal of Agricultural Research voi. xxn. no. s 



referred to below were prepared as follows : Living worms were removed 

 from the intestine of a calf, washed a number of times in physiological 

 salt solution, and kept in a refrigerator at a temperature of 8° C. over- 

 night. The following day the worms which were still alive were trans- 

 ferred to fresh salt solution and crushed in a mortar. The crushed 

 material was then suspended in about two volumes of physiological salt 

 solution, shaken thoroughly, and centrifuged. The supernatant fluid 

 which was opalescent was removed and tested as to its hemolytic power. 

 Tested on a 3 per cent suspension of washed sheep blood corpuscles, it 

 was found that 5 drops of the extract hemolyzed 5 drops of the suspen- 

 sion of blood corpuscles in i hour at a temperature of 37°. Even i 

 drop of extract hemolyzed 5 drops of the suspension of corpuscles after 

 a few hours. Controls, that is, 5 drops of suspension of corpuscles plus 

 5 drops of salt solution, remained intact. It was observed that before 

 hemolysis set in the contents of the tubes assumed a dark red hue. 



An extract from another lot of Bustomum phlehoiomum prepared as has 

 already been described was tested on a 5 per cent suspension of washed 

 rabbit cells. Five drops of extract produced hemolysis rather slowly 

 upon 5 drops of suspension of corpuscles. 



In a third experiment an extract prepared from worms that had been 

 kept in a refrigerator overnight was tested on four different tubes of cat- 

 tle erythrocytes and on four different tubes of hog erythrocytes. The 

 extract in question was prepared from living specimens as follows: 

 Forty-five specimens were ground up in a mortar and suspended in 2 cc. 

 of physiological salt solution. The suspension was centrifuged, and the 

 opalescent fluid was removed and tested on a 5 per cent suspension of 

 washed blood corpuscles at 37° C. Three drops of extract were added 

 to 5 drops of corpuscle suspension. The experiments with each sample 

 of corpuscle suspension were controlled by adding 3 drops of physiological 

 salt solution to 5 drops of suspension of blood cells. The results of these 

 experiments follow. 



Cattle blood corpuscles. — ^After i hour one tube of blood was par- 

 tially hemolyzed and three tubes were intact. After 2% hours two tubes 

 were completely hemolyzed and two were intact. After 3 hours three 

 tubes were hemolyzed ; one was intact. The tubes containing the mixtures 

 were placed in a refrigerator at 8° C. until the next day. When examined 

 hemolysis was complete in all tubes. The controls showed no hemolysis. 



Hog blood corpuscles. — ^After i hour all tubes were intact. After 

 1^4 hours two tubes were partially hemolyzed and two intact. After 2}^ 

 hours two tubes were partially hemolyzed and two completely hemolyzed. 

 After 3 hours all tubes showed complete hemolysis. Controls were intact. 



Inasmuch as it was found that the hemolysin could be preserved by 

 drying the worms, powdering the dried material, and storing it in a dark 

 place, further experiments with Bustomum phlebotomum hemolysin were 

 performed with dried material. The details of these experiments follow. 



