DETECTION AND IDENTIFICATION 59 



and glacial acetic acid about 4 drops. The first two can 

 l)e kept mixed without deterioration, but the glacial acetic 

 acid should be added just before fixation. Fix at room 

 temperature. The fixative is put in a Petri (preferably 

 square) dish or a wide stendor dish. The smear is gentlv 

 dropped on the fixati\e with the smeared surface facing 

 downward. With a little practice, air bubbles can be 

 a\'oided and make the smear float on the surface of the 

 fixative. After about one minute, turn the smear around 

 and let it stav on the bottom of the container for 5 more 

 minutes. If the smear is too thick, it will not float on the 

 fixation. A thin coat of \ aseline on the unsmeared side will 

 allow the co\ er to float. About six co\'er-glass smears can 

 be fixed in an ordinarv Petri dish simultaneously. 



The cover-glasses are now transferred into a coplin jar 

 or better Columbia staining jar for cover-glasses, containing 

 50% alcohol for 10 minutes, and then two changes for 10 

 minutes each. Transfer next the smears into a jar containing 

 30^ alcohol for 5 minutes, and then into a jar with water 

 which is now placed under gentlv running tap water for 

 15 minutes. Rinse the smears in distilled water. They are 

 ready for staining. 



The most dependable staining which is wideh^ used for 

 staining intestinal protozoa is Heidenhain's iron haema- 

 toxylin. It requires a mordant, ammonio-ferric sulphate (iron 

 alum) and a dve, haematoxylin. Crystals of iron alum be- 

 come vellow and opaque verv easily; select clear \iolet 

 crvstals and prepare 2% waterv solution which will keep 

 for a long time. 0.5-1^/c haematoxvlin must be well "ripe." 

 The most convenient way of preparing it is to make 10% 

 absolute alcohol solution. B\ diluting this stock solution 



