122 Methods in Plant Histology 



material is hard, like many woody stems, it will cut better after 

 remaining in this mixture for a couple of weeks. 



The following schedules, beginning with the colloidin sections 

 in 70 per cent alcohol, will give the student a start in the staining: 



Delafield's Haematoxylin and Eosin. 



1. 70 per cent alcohol, 2 to 5 minutes. 



2. Delafield's haematoxylin, 5 to 30 minutes. 



3. Wash in water, 5 minutes. 



4. Acid alcohol (1 c.c. hydrochloric acid+100 c.c. of 70 per cent alcohol) 

 until the stain is extracted from the celloidin, or at least until only a 

 faint pinkish color remains. 



5. Wash in 70 per cent alcohol (not acid) until the purple color returns. 



6. Stain in eosin (preferably a 1 per cent solution in 70 per cent alcohol), 

 2 to 5 minutes. 



7. Dehydrate in 95 per cent alcohol, 2 to 5 minutes. Do not use abso- 

 lute alcohol unless you wish to dissolve the celloidin, which is not 

 necessary with this staining. 



8. Clear in Eycleshymer's clearing fluid, usually 1 to 2 minutes, but 

 sometimes 5 to 10 minutes. 



9. Mount in balsam. 



Safranin and Delafield's Haematoxylin .- 



1. 70 per cent alcohol, 2 to 5 minutes. 



2. Safranin (alcoholic), 6 to 24 hours. 



3. Acid alcohol (a few drops of hydrochloric acid in 70 per cent alcohol) 

 until the safranin is removed from the cellulose walls. 



4. Wash in 50 per cent alcohol, 5 to 10 minutes to remove the acid. 



5. Delafield's haematoxylin, 2 to 5 minutes. 



6. Wash in water, 5 minutes. 



7. Acid alcohol, a few seconds. 



8. Dehydrate in 95 per cent alcohol, 2 to 5 minutes, then in absolute 

 alcohol, 2 to 5 minutes, which will partially dissolve the celloidin. 



9. Clear in clove oil, which will complete the removal of the celloidin. 

 10. Be sure that the sections are free from fragments of celloidin and then 



mount in balsam. 



Jeffrey's improvements in the celloidin method have been 

 described in considerable detail by Plowman. 1 Sections of hard 

 stems and roots cut by this method could hardly be surpassed, 



1 Plowman, A. B., The Celloidin Method with Hard Tissues, Botanical Gazette, 37: 

 456-461, 1904. 



