COMPLEMENT FIXATION IN GONOCOCCUS INFECTIONS 507 



If the antigen is new and the anticomplementary dose is entirely 

 unknown, it may be necessary, in making this titration, to use a different 

 dilution, with higher and lower doses. In conducting the main test the 

 foregoing antigen could be used in dose of 0.2 or 0.4 c.c. of this dilution. 



The Test. The serums should be fresh and clear, and heated to 56 C. 

 for one-half hour. For each serum use four test-tubes (12 by 1 cm.), 

 arranged in a row. Into each of the first three place the dose of antigen 

 and increasing doses of serum 0.05 c.c., 0.1 c.c., 0.2 c.c.; the fourth 

 tube is the serum control, and into this is placed the maximum dose of 

 serum (0.2 c.c.) but no antigen; 1 c.c. of complement diluted 1 : 20 is 

 added to each tube. The following controls are included : 



1. A positive control with an antigonococcus serum or with the serum 

 of a patient who reacted positively on a former occasion. 



2. A negative control with the serum of a healthy person. 



Both of these controls may be set up with but the maximum dose of 

 serum (0.2 c.c.). 



3. The serum control of each serum is conducted in the fourth tube of 

 each series. At the completion of the test this tube should show com- 

 plete hemolysis and thereby indicate that the serum was not anticom- 

 plementary. 



4. The antigen control at this time includes the dose of antigen and 

 complement. 



5. The hemolytic system control at this time receives the dose of 

 complement. 



6. The corpuscle control receives 1 c.c. of the corpuscle suspension. 

 To each tube sufficient saline solution is added to bring the total 



volume up to about 2 c.c. The tubes are shaken and incubated for one 

 hour at 37 C. in the thermostat or in a water-bath (not less than one 

 hour), when 2 units of antisheep amboceptor and 1 c.c. of sheep corpuscle 

 suspension are added to each tube except the corpuscle control. The 

 tubes are gently shaken again and reincubated for an hour or longer, 

 depending upon the hemolysis of the controls, after which the results 

 are recorded. This secondary incubation may be omitted and the tubes 

 placed in a refrigerator overnight and the results read the next morning. 

 Under these conditions hemolysis occurs slowly, and, according to some 

 workers in this field, the reaction becomes more delicate. 



Conducting the primary incubation by placing the tubes in a refrigerator 

 at8C. for four hours or over night after the method of McNeal, followed by 

 the addition of 2 units of hemolysin and 1 c.c. of corpuscle suspension to 

 each tube and incubation in a water-bath at 38 C. for one-half to one hour, 



