AMBOCEPTORS AND COMPLEMENTS 931 



EXERCISE 33. AMBOCEPTORS AND COMPLEMENTS (Continued) 

 EXPERIMENT 84. TITRATION OF HEMOLYTIC COMPLEMENT 



1. Prepare 20 c.c. of complement 1:20; prepare a 2^ per cent, suspension of 

 sheep cells. 



2. To a series of 10 test-tubes add increasing doses of diluted complement serum: 

 0.1, 0.2, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, and 1.5 c.c.; add 1^ units of hemolytic ambo- 

 ceptor (determined by previous titration) ; 1 c.c. of corpuscle suspension and sufficient 

 salt solution to bring the total volume in each tube to 3 c.c. 



3. Shake gently and incubate for one hour at 37 C. 



(a) Record the results. What is the complement unit of this serum? 



(b) What animal serum is best adapted for complement in hemolytic 

 work? 



(c) Is the complement content of the serums of different animals 

 constant? Does it vary in animals of the same species? In the same 

 animal at different times? 



EXPERIMENT 85. PHENOMENON OF COMPLEMENT FIXATION 



This experiment is introduced here to show the Bordet-Gengou 

 phenomenon of complement fixation. The exact technic of complement- 

 fixation reactions as conducted for the diagnosis of syphilis and other 

 infections will be given in subsequent exercises. 



1. Use the same complement serum, amboceptor and corpuscle suspension as 

 used in the preceding experiment. The unit of complement is now known. 



2. Secure 1 c.c. of an antigonococcus and a normal serum and heat at 56 C. for 

 thirty minutes. 



3. Secure an emulsion of gonococci which is now called the antigen: for the proper 

 dose to use consult the instructor. 



4. Proceed as follows: 



Tube 1 : 0.2 c.c. antigonococcus serum + dose of antigen + 2 units of comple- 

 ment + normal salt solution. 



Tube 2: 0.2 c.c. antigonococcus serum + 2 units of complement + normal salt 

 solution (serum control). 



Tube 3: 0.2 c.c. normal serum + doses of antigen + 2 units of complement -f 

 normal salt solution. 



Tube 4: 0.2 c.c. normal serum + 2 units of complement -f normal salt solution 

 (serum control). 



Tube 5 : dose of antigen + 2 units of complement + normal salt solution (anti- 

 gen control). 



Tube 6: 2 units of complement + normal salt solution (hemolytic control). 



Tube 7: 1 c.c. corpuscle suspension + normal salt solution (corpuscle control). 



Plug tube with cotton as it is finished. 



5. Shake all tubes gently and incubate for one hour at 37 C. 



6. Add 1% units of amboceptor and 1 c.c. corpuscle suspension to all tubes 

 except corpuscle control. Shake gently and reincubate for one hour. 



