416 THE TECHN1C OF COMPLEMENT-FIXATION REACTIONS 



ligible. At any rate it is very simple and but little trouble to remove the 

 antisheep amboceptor routinely from human serums previous to mak- 

 ing the tests) (for technic, see p. 378). 



A method for titrating immune amboceptor has been given on p. 375. 

 This important feature will be dealt with again in giving a detailed 

 description of the various methods that follow. 



IV. Red Blood Corpuscles. Defibrinated sheep blood is washed 

 three times with an excess of sterile normal salt solution to remove all 

 traces of serum (p. 28). 



In the original Wassermann reaction a 5 per cent, suspension in salt 

 solution is used in doses of 1 c.c. This emulsion is quite heavy, and 

 sharper and clear results are secured by using just half this amount and 

 at the same time sufficient cells are used to make the readings easy and 

 distinct. Either 0.5 c.c. of a 5 per cent, or 1 c.c. of a 2.5 per cent, sus- 

 pension may be used. I have used the latter with entire satisfaction for 

 several years. 



The emulsion of cells is prepared with sterile 0.85 per cent, sodium 

 chlorid solution. To 2.5 c.c. of corpuscles sufficient salt solution is 

 added to bring the total volume of the emulsion up to 100 c.c., or smaller 

 amounts may be prepared by suspending 1 c.c. of corpuscles in 39 c.c. 

 of salt solution. 



Before each day's work the amount of corpuscle suspension needed 

 should be calculated, and sufficient for the day prepared at one time, for 

 if a fresh emulsion is prepared later, titration with the complement and 

 amboceptor would be required. Attempts to count the corpuscles in 

 suspension can only be regarded as approximate and are unreliable. 

 By titrating each emulsion with the complement and amboceptor to be used, 

 that particular emulsion is thereby adjusted, so that it is immaterial whether 

 a few more or a few less corpuscles are present. 



Sheep's blood is obtained either from an abattoir or by bleeding an 

 animal from the external jugular vein (p. 48).. The latter method is 

 preferable, but due care must be exercised not to bleed too frequently 

 or in excessive amounts, as if anemia occurs the corpuscles become 

 unduly fragile. 



Sheep's cells are easily preserved in a satisfactory condition for 

 forty-eight hours by first washing them and then storing the sedimented 

 corpuscles in a good ice-chest. Suspensions are less easily preserved. 

 It is best to use fresh corpuscles, and those that are several days old 

 and unduly fragile should never be used. Attempts to preserve cor- 

 puscles with the aid of mercury bichlorid, formalin, etc., have not yielded 

 satisfactory results. 



