STAIN TECHNIQUE 19 



Selecting a combination of stains. Test the object with a 

 stain from each of the four classes : acid, direct, mordant, and 

 basic. If the best colors are given with basic and mordant stains, 

 the combination of Safranine and Haematoxylin may be tried. 

 Should the direct dye work better than Haematoxylin, use 

 Safranine and Niagara Sky Blue. Many sections of growing tis- 

 sues fail to differentiate with a combination containing a basic dye. 

 If both the acid and direct stains provide clear colors on the test 

 material, the desired contrast is obtained by using either an acid 

 and a direct dye or two acid dyes of contrasting colors. The direct 

 dyes may be used as contrast stains with either an acid or a basic 

 dye. 



Combinations of two stains. 



Rule : Use the lighter stain first. 

 Example No. 1 — Crocein Scarlet MOO, C. I. No. 252, and Niagara 



Sky Blue, C. I. No. 520. 



This red acid dye and blue direct dye provide an adequate con- 

 trast on small invertebrates, paraffin sections of animal tissues, 

 meristems, or on fungi. The combination of stains may be used in 

 two ways, either separately or mixed in solution. 

 I. Two dyes used separately. 



1. Ten minutes in Crocein Scarlet MOO. 



2. Rinse the material in distilled water. 



3. One minute in Niagara Sky Blue. 



4. Rinse the material in distilled water. 



5. Objects used as whole mounts are given three minutes 

 in each grade of alcohol: 35, 50, 70, 85, 95, and abso- 

 lute alcohol. Paraffin sections are transferred from 

 water to 95 per cent alcohol and then into absolute 

 alcohol, one minute in each. 



6. Materials used as whole mounts need three minutes 

 in each mixture: \ xylene, \ xylene, J xylene and 

 xylene. Paraffin sections are given one, two, three, 

 and five minutes respectively in the alcohol-xylene 

 mixtures. 



7. Whole mounts are infiltrated gradually in balsam. 



