24 MATERIALS AND PROCESSES OF SLIDE-MAKING 



3. Give each slide an individual quick rinse in tap water and pass it directly 

 to 96 per cent alcohol. 



4. Keep the slides in motion in 96 per cent alcohol until they are dehydrated. 



5. Then rinse each section once or twice in absolute alcohol and transfer 

 it to xylene. 



6. As soon as the section is clear, mount it in balsam. 



This stain relies for its effect on the fact that muscular tissues retain the 

 yellow color of the picric acid more readily than the other connective tissues. 

 It is the best known of the double contrast solutions but has certain serious 

 disadvantages. In the first place, picric acid tends to remove hematoxylin from 

 the nuclei, so that one must overstain considerably in the hematoxylin if one 

 is to secure adequately blue nuclei in the finished mount. The second major 

 objection is that picric acid is soluble to a certain extent in xylene so that the 

 yellow color is gradually extracted. Sections stained by this method and mounted 

 in balsam are rarely of very much use a year after they have been prepared. To 

 avoid these objections, the author much prefers: 



Gray's Double Contrast Stain: 

 Staining solution 



Water 100 ml. 



Orange II 0.6 Gm. 



Ponceau 2R 0.4 Gm. 



SOLUTION IS USED AS FOLLOWS: 



1. Accumulate sections, with nuclei stained either in hematoxylin or in 

 celestin blue B, in tap water. 



2. Transfer the slides to the staining solution for 2 minutes. 



3. Remove each slide individually from the stain, drain, blot, and then dip 

 up and down in absolute alcohol until sufficiently differentiated. The com- 

 pletion of differentiation and the completion of dehydration usually coincide. 



4. Then transfer each slide to xylene and mount in the ordinary manner. 



This stain gives a good range of red-orange and gold shades on most histo- 

 logical sections and is no more difficult to use than a simple solution of eosin. 



Double contrast stains for sections in which the nuclei have been stained red 

 are not very common, though the following, originally designed for use with 

 heavily yolked embryonic material, is really excellent. 



Smith's Picro-Spirit Blue: 

 Staining solution 



Absolute alcohol 100 ml. 



Picric acid 1 Gm. 



Spirit blue Enough to saturate 



