212 ESSENTIALS OF BACTERIOLOGY 



sterile test-tubes. After clotting has taken place the clear 

 serum is removed. This is the amboceptor serum. 



Washed sheep corpuscles are obtained by centrifuging de- 

 fibrinated sheep blood, pipeting off the serum, replacing it 

 with normal salt solution, shaking, and again centrifuging. 

 This is repeated three times. 



Patient's serum obtained from blood from the patient's arm 

 is heated thirty minutes at 56 C. to destroy complement. 



Titration or Testing of Reagents. Titrate amboceptor. 

 One c.c. of a 5 per cent, suspension of washed sheep cor- 

 puscles in salt solution and o.i c.c. of fresh guinea-pig 

 serum are added to a series of test-tubes. The amboceptor 

 serum is then added so that each tube receives more than the 

 preceding one. Salt solution is added to make 5 c.c. and 

 the tubes incubated for two hours at 37 C. with occasional 

 shaking. That tube in which complete hemolysis has taken 

 place in just two hours contains ^ unit of amboceptor. 



Titration of Complement. Into each of a series of tubes 

 place i c.c. of the corpuscle suspension and ^ unit of ambo- 

 ceptor. Next add 0.6, 0.7, 0.8, 0.9, i, i.i, 1.2 c.c. of fresh 

 guinea-pig serum respectively and incubate for two hours, 

 shaking occasionally. Those tubes which show complete 

 hemolysis in just two hours contain i unit of complement. 



Titration of Antigen. Two- tenths c.c. of serum, pre- 

 viously heated to 56 C. for a half-hour, from a known, un- 

 treated case of secondary syphilis, and i unit of complement 

 are added to each of a series of test-tubes. Antigen is now 

 added, so that each tube contains more than the preceding 

 one, and salt solution added and brought to 3 c.c. The 

 mixture is incubated for one hour at 37 C., at the end of 

 which time 2 units of amboceptor and i c.c. of corpuscle 

 suspension are added and the tubes returned to the incubator. 

 After a short period the tube containing the smallest amount 

 of antigen will show complete hemolysis. As the dose of 

 antigen is increased the amount of hemolysis is decreased 

 until a point is reached at which no hemolysis takes place 

 even after twenty-four hours. The first tube in the series 



