SERUM DIAGNOSIS OF PAROXYSMAL HEMOGLOBINURIA 381 



make 500 c.c. of salt solution, ranging from 0.1 to 0.6 per cent, in grada- 

 tions of 0.02 per cent. This means the preparation of twenty-six dif- 

 ferent solutions, which should be preserved in proper-sized bottles fitted 

 with tight rubber stoppers. 



When the test is needed only occasionally, these solutions are readily 

 prepared by filling a 50 c.c. buret, graduated in one-tenths, with dis- 

 tilled water, and another with a 1 per cent, solution of pure dried sodium 

 chlorid. From these, the various solutions are readily prepared after 

 the following manner: 



10 c.c. of 0.6 per cent, sodium chlorid = 6 c.c. of 1 per cent, salt 



solution + 4 c.c. of 

 distilled water. 



10 c.c. of 0.58 per cent, sodium chlorid = 5. 8 c.c. of 1 per cent, salt 



solution + 4.2 c.c. of 

 distilled water. 



10 c.c. of 0.56 per cent, sodium chlorid = 5.6 c.c. of 1 per cent, salt 



solution + 4.4 c.c. of 

 distilled water. 



10 c.c. of 0.54 per cent, sodium chlorid = 5.4 c.c. of 1 per cent, salt 



solution -h 4.6 c.c. of 

 distilled water. 



10 c.c. of 0.52 per cent, sodium chlorid = 5. 2 c.c. of 1 per cent, salt 



solution -f 4.8 c.c. of 

 distilled water. 



10 c.c. of 0.5 per cent, sodium chlorid = 5 c.c. of 1 per cent, salt 



solution -f- 5 c.c. of 

 distilled water. 



Similar dilutions are made, until the final dilution is reached. In 

 many instances it may not be necessary to use so large a number of dilu- 

 tions, as from 0.5 to 0.2 per cent, may be sufficient range to indicate the 

 tonicity. 



Five cubic centimeters of blood are aspirated, under aseptic pre- 

 cautions, from an arm vein of the patient, and immediately placed in 

 25 c.c. of sterile 1 per cent, sodium citrate in 0.85 per cent, sodium chlorid 

 to prevent coagulation. The flask or large centrifuge tube is well 

 shaken, and the mixture is centrif uged at sufficient speed to throw down 

 the corpuscles. The supernatant fluid is drawn off, and the corpuscles 

 are washed once or twice more with sterile normal salt solution. After 

 the last washing the supernatant fluid is removed, leaving the erythro- 

 cytes at the bottom of the tube. 



