AMBOCEPTORS AND COMPLEMENTS 851 



EXPERIMENT 76. GENERAL PROPERTIES OF AMBOCEPTORS 



1. Secure antisheep amboceptor so diluted that 1 c.c. contains 1 hemolytic unit. 



2. Prepare a 2^ per cent, suspension of sheep corpuscles. 



3. Secure fresh guinea-pig complement serum and dilute 1:20. 



4. Place two units of amboceptor into each of six test-tubes. Place these in a 

 water-bath and heat at 65 C. 



5. Remove a tube from the water-bath after fifteen minutes, thirty minutes, 

 forty-five minutes, one hour, one and one-half hours, and two hours. After allowing 

 them to cool, add 1 c.c. corpuscle suspension and 1 c.c. complement serum (1:20). 



6. Set up a tube containing two units of amboceptor (not heated) + 1 c.c. cor- 

 puscle suspension +1 c.c. complement serum (control). Shake each tube gently and 

 incubate for an hour at 37 C. 



(a) Has the amboceptor deteriorated by exposure to this degree of 

 heat? 



(b) Discuss the stability of amboceptors to temperature, age, germi- 

 cides, and drying. 



(c) Discuss the relative stability of amboceptors and complements. 



EXERCISE 30. AMBOCEPTORS AND COMPLEMENTS. HEMOLYSINS 



(Continued) 



EXPERIMENT 77. MECHANISM OF AMBOCEPTOR ACTION 



1. Secure antisheep amboceptor so diluted that 1 c.c. contains one hemolytic 

 unit. 



2. Prepare a 2 1 A per cent, suspension of sheep corpuscles. 



3. Secure fresh guinea-pig serum and dilute 1:20. 



4. Into two centrifuge tubes place two units of amboceptor and 1 c.c. of cor- 

 puscle suspension. Mix, and place one tube in a glass of cracked ice, leaving the 

 other at room temperature. After one hour centrifuge both. Pipet the supernatant 

 fluids into two separate test-tubes. 



5. To the sedimented corpuscles in both centrifuge tubes add 1 c.c. corpuscle 

 suspension and 1 c.c. diluted complement serum. Mix well. Incubate tubes for 

 one hour at 37 C. 



6. To the supernatant fluid of each tube add 1 c.c. corpuscle suspension and 1 

 c.c. of diluted complement serum. Mix. Incubate tubes for an hour. 



(a) In which tubes has hemolysis occurred? 



(b) How do you explain the results? 



(c) Discuss the prevailing views of amboceptor action. 



EXPERIMENT 78. A FURTHER STUDY OF THE MECHANISM OF AMBO- 

 CEPTORS 



1. In a centrifuge tube place six hemolytic doses of antisheep hemolysin and 2 c.c. 

 of fresh guinea-pig complement diluted 1 : 20. Place in a glass of cracked ice until the 

 mixture is thoroughly chilled. Then add 2 c.c. of a 2^ per cent, suspension of sheep 

 cells (also chilled). Mix and keep at a low temperature for an hour, centrifuge 

 thoroughly, and pipet the supernatant fluid to a separate test-tube. 



