STANDARD METHODS AND REAGENTS 465 



Simple sedimentation in a tall cylinder is commonly employed 

 for the detection of all cyst-producing forms. The technique may 

 be refined as described in Chapter XXXI (p. 281). 



The following method, adapted from Yorke and Adams (1926), 

 was devised for use with Endani^ba histolytica but it would prob- 

 ably be equally useful for the other intestinal amoebae and flagellates 

 that produce cysts. The specific gravity of the sugar solution is not 

 sufficiently high to carry coccidial oocysts to the surface in any 

 great quantity. One of the advantages of this method is that the 

 cysts are recovered in a viable state and can be used for inoculation, 

 culture, and various other purposes. Procedure is as follows : 



A mass of feces is mixed with a little water until it is thoroughly emul- 

 sified. Shake the emulsion with from 500 to 1000 c.c. of water, pour into a 

 tall glass cylinder and allow it to stand about fifteen minutes. Skim off the 

 flotage with a wire loop covered with cheesecloth. Pour off the supernatant 

 fluid and save it. Discard the lowest inch with the sediment. 



Either centrifuge the supernate or allow it to stand overnight in a tall 

 glass cylinder. In either case the sediment is shaken up thoroughly with a 

 solution of cane sugar in water, of a specific gravity of about 1080 (20 

 grams of sugar in 100 c.c. of solution) and is centrifuged at high speed for 

 two or three minutes. The upper one-fifth of the supernate is pipetted into 

 a clean centrifuge tube, and the volume is made up to nearly capacity with 

 water and is again centrifuged at high speed. The resulting sediment is 

 examined for cysts. 



The next method, adapted from Boeck's (1917) modification of 

 the Cropper and Row (191 7) technique gives satisfactory results 

 with any kind of intestinal protozoan cyst. It is as follows : 



At least I gram of feces is placed in 30 c.c. of normal saline and is stirred 

 with the soda fountain mixer (or is thoroughly emulsified by some other 

 more convenient method such as shaking with glass beads or shot) for from 

 2 to 10 minutes depending upon the consistency of the stool. When the 

 specimen appears to be uniformly emulsified, 5 c.c. 01 ether is added and 

 the resulting mixture is stirred or shaken for an additional 2 to 3 minutes. 



Caution: i. If obliged to shake the ether mixture in a closed vessel, 

 be careful to reduce the internal pressure at frequent intervals by 

 opening the container ! 



2. Keep away from any open flame while shaking this mixture or 

 releasing the gases from it ! 



The emulsion should be quickly placed in a separatory funnel, and allowed 

 to stand for from 5 to 7 minutes. (Unless the soda fountain mixer is used 

 to make the ether mixture, the mixing can be most easily accomplished in 

 the separatory funnel.) After the required time, about 15 c.c. of the material 

 is withdrawn from the bottom of the separatory funnel into a centrifuge 



