TECHNIQUE 471 



TECHNIQUE. 



PRESERVATION AND EXAMINATION OF FLUKES. 



Fixation. (Method A.) (i) Place the flukes in a test tube or small bottle 

 a quarter full of normal saline. Shake the contents as hard as possible (the 

 object of this is to extend the flukes) for half a minute. 



(2) Add immediately an equal bulk of saturated aqueous solution of corro- 

 sive sublimate and shake again as vigorously as possible for a few minutes. 



(3) Transfer when convenient to 70 per cent, alcohol. (Before staining 

 and mounting remove the sublimate with tincture of iodine.) 



(Method B.) In case of large flukes, e.g., Fasciola hepatica, Fasciolopsis 

 buski, compress the flukes between two glass slides with rubber bands or 

 thread. Fix in sublimate or in absolute alcohol, or in 10 per cent, formalin. 



(Method c.) Place the flukes in 10 per cent, formalin solution. 



Staining is successfully effected by using quite dilute solutions of carmine 

 or haematein overnight. This is far preferable to using strong solutions, 

 as it may be almost impossible to remove a too intense stain. Almost any 

 dilute carmine solution suffices. One of the best is acetic-alum carmine 

 (boil excess of carmine in a saturated aqueous solution of potash-alum for 

 about fifteen minutes ; add glacial acetic acid to the extent of 10 per cent. ; 

 let it stand for a week ; filter). For use, dilute about thirty times with 

 water. Place the flukes directly in the stain. Stain overnight or longer. 



Differentiation. In order to get the sharpest picture, it is best now to 

 differentiate (but this may often be omitted) with acid alcohol (70 per cent, 

 alcohol 100 parts, HC1 5 drops). Allow to act from one to twenty-four 

 hours, according to the appearance of the flukes. Similarly, in staining with 

 hamatoxylin solution, dilute twenty to thirty times so that the water is merely 

 tinged with the stain. Differentiate as before. After staining, dehydrate, 

 clear, and mount in balsam if required. 



Clearing and Mounting. (i) Carbolic acid (carbolic acid 94, water 6) is 

 a very convenient clearing agent. It may be used for stained or unstained 

 specimens. It will clear rapidly without previous dehydration. If it is 

 required to mount a specimen permanently, transfer from carbolic to 

 alcohol, then cedar-wood oil (or xylol, etc.), then balsam. 



(2) Creasnte. Dehydrate the specimen, stained or unstained, transfer to 

 creasote. If it is desired to mount permanently, transfer back to alcohol, 

 then cedar-wood oil, then balsam'. 



(3) Cedar-wood Oil. Preferable to xylol or oil of cloves. Dehydrate 

 the specimen in alcohol. To mount permanently, transfer to balsam. 



