390 Special Procedures II (chap. 23) 



glass. Drop immediately into iodine-alcohol (page 12). If smears 

 wash off, too much PVA was carried over; leave material for longer 

 time on cleansing tissue. 



Smears may be fixed directly on slides. Take 1 drop of fecal material 

 to 3 drops of PVA fixative. Smear over a large area of the slide and 

 dry in oven overnight. Immerse in iodine-alcohol. 



1. Leave in iodine-alcohol: 1 minute. 



2. Decolorize in 70% alcohol, 2 changes: 1-2 minutes each. 



3. Stain: 5-10 minutes. 



4. Differentiate in acidified 90% alcohol (1 drop glacial acetic acid/ 

 10 ml. alcohol): 10-20 seconds or until stain no longer runs from 

 smear. 



4. Dehydrate in absolute alcohol; rinse twice, dipping up and down. 



6. Dehydrate in second change of absolute alcohol: 1 minute. 



7. Clear and mount. 



results: 



background — predominantly green 



cysts — bluish-green cytoplasm, purplish-red nuclei; green is more in- 

 tense than background stain 

 engulfed red cells — vary, either green, red, or black 

 chromatoidal bodies — intense green, but may be reddish 



comments: 



If cysts do not stain or stain predominantly red, fixation is incom- 

 plete. Warming the fixative may help, although cold fixation yields 

 more critical staining. A newly prepared stain is predominantly red 

 in color and reaction; older stains show more violets and greens. 



Warjiing: Press the cover glass in place very gently; the smear is 

 somewhat brittle and is easily loosened. Too much pressure on the 

 cover glass may start parts of the smear to floating around on the 

 slide. 



This stain does fade after a few years. 



References: California State Department of Public Health, Bacteriol- 

 ogy Department, Division of Laboratories, Berkeley, California; Go- 

 mori (1950) and Wheatley (1951) 



Amoebae in Tissue 



Meriweather (1934) Method 



fixation: 10% formalin or a mercuric chloride fixative. 



