84 The Preparation of Microscope Slides 



Differentiating solution 



0.1% hydrochloric acid in 70% alcohol 

 Bluing solution 



0.1% sodium bicarbonate in tap water 



Method for Sections 



1. Accumulate slides in tap water. 



2. Stain until sections are dark purple. About 5 min for animal tissues 

 or several hours for plant sections. 



3. Rinse in distilled water. 



4. Differentiate until nuclei or xylem only remain colored. 



5. "Blue" until nuclei or xylem dark blue, usually from 1 to 2 min. 



Indirect Hematoxylin Staining. Indirect staining is that method in 

 which very dilute stain is applied from a solution containing a mordant. 

 The stain builds up slowly on the nuclei but is not absorbed by the 

 cytoplasm. This method is never used on sections but is excellent for 

 wholemounts. Any alum hematoxylin may be used by the following 

 method: 



Staining solution 



Delafield's hematoxylin 1 ml 



1% ammonium alum 20 to 100 ml 



Note: The larger the object the more dilute must be the stain. A proto- 

 zoan can be adequately stained in a 1 : 20 dilution. A liver fluke would 

 require at least a 1 : 100 dilution. 



Washing solution 



1% ammonium alum 

 Bluing solution 



0.1% sodium bicarbonate 



Method for Wholemounts 



1. Accumulate fixed and well-washed objects in distilled water. 



2. Place in stain until sufficiently stained, that is, until internal struc- 

 tures are clearly seen. 



3. Wash in washing solution until no more color comes away. 



4. Blue in bluing solution until all dye is converted to blue form. This 

 requires at least 5 min for a small hydroid and as much as 12 hr for a 

 large liver fluke. Lack of adequate bluing is responsible for most failures 

 with this technique. 



CARMINE 



This substance is a compound of carminic acid, derived from the 

 cochineal insect, with tin and aluminum compounds. The dye is not 



