178 METHODS FOR THE ANALYSIS OF BLOOD 



added to the flask, the whole is made to volume, mixed, and let 

 stand with occasional shaking for at least ten minutes to allow 

 laking of the corpuscles. 5 c.c. of this dilution are run slowly, 

 with shaking, into 15 to 18 c.c. of the acid ammonium sulfate in 

 a 25 c.c. flask, the volume is made to 25 c.c. with the ammonium 

 sulfate solution, and the whole well mixed. Of this mixture 10 

 c.c. are measured out and used for the determination of acid-solu- 

 ble phosphoric acid as directed below. The remainder is allowed 

 to stand at least ten minutes, then filtered, and determinations 

 are made of inorganic phosphates. 



Inorganic. 5 c.c. of the filtrate ( = 0.5 c.c. of corpuscles) are 

 used for the determination which is carried out as in method (6) 

 for plasma, using the same standard. Determinations should be 

 made at once after filtering since in certain cases and particularly 

 in warm weather the values have been found to increase on stand- 

 ing, probably at the expense of the unknown acid-soluble phos- 

 phoric acid compound. 



Acid-Soluble. 10 c.c. of the ammonium sulfate-.corpuscle 

 mixture are measured into a small flask and immersed for two 

 minutes in boiling water. After cooling it is poured into a 25 c.c. 

 graduated flask, the small flask is rinsed with 10 c.c. of the acid 

 ammonium sulfate, and the rinsings are added to the main portion 

 in the 25 c.c. flask. The volume is made up to the mark with 

 water, mixed, and filtered. For the determination 2 c.c. of the 

 filtrate (0.08 c.c. of corpuscles) are measured into one of the large 

 test-tubes, glass beads and 1.5 c.c. of the sulf uric-nitric acid 

 mixture are added, the whole is digested, and determinations are 

 made as with total phosphates, the small amount of ammonium 

 sulfate present not interfering. For the standard, 5 c.c. of the 

 standard phosphate (0.15 mg. of HsPC^) are used and to it are 

 added 1.5 c.c. of the ammonium sulfate solution; then it is treated 

 with alkali, neutralized, etc., as in the regular determinations: 



Notes on the Methods 



Solutions Required for these Determinations. 1. The strych- 

 nine molybdate reagent as described. 



2. Add ammonium sulfate. Saturated ammonium sulfate, 

 free from all but traces of phosphate, to which have been added 

 15 c.c. of glacial acetic acid per liter. 



