94 METHODS IN PLANT HISTOLOGY 



5. Xylol: 2 or 3 c.c. of absolute alcohol may be added to each 100 c.c. of 

 xylol, if the free light green shows a tendency to precipitate, 



6. Mount in balsam. 



This stain is particularly good for phloem. Since the light green is 

 not likely to overstain and only slightly weakens the safranin, the 

 combination is a rather easy one, so that even the beginner can hardly 

 fail to get a good preparation. 



Malachite green and Congo red. — I am indebted to Dr. Sharp for 

 this method, which has been popular in Professor Gregoire's labora- 

 tory at Louvain. 



Sections of fresh material should be treated with 95 per cent alco- 

 hol and then transferred to water. 



1. Three per cent aqueous solution of malachite green or methylene blue, 

 G hours or more. 



2. Wash in water. 



3. Congo red, 1 per cent aqueous solution, 15 minutes. 



4. Wash in water. 



5. Rinse in 80 per cent alcohol. As soon as the malachite green or anilin 

 blue color appears through the red, transfer quickly to 



6. Absolute alcohol. 



7. Xylol. 



8. Balsam. 



Iodine green and acid fuchsin is another good combination for such 

 sections. The stain will be particularly brilliant if sections from fresh 

 material are fixed in 1 per cent chromo-acetic acid for 24 hours; and 

 then washed for an hour in water. Beginning with the sections in 

 water, the procedure is as follows : 



Stain in aqueous iodine green for 12-24 hours. Then wash in water 

 until the stain is nearly all washed Out from the cellulose walls but is 

 still brilliant in the lignified walls. If the stain acts for too short a 

 time, the washing-out process necessary to remove the stain from the 

 cellulose walls will leave only a pale green color in the lignified walls. 

 Stain in aqueous acid fuchsin for 2-10 minutes. This should stain the 

 cellulose walls sharply, but should not act long enough to affect the 

 lignified tissues. Pour off the stain (which may be used repeatedly), 

 and pour on 95 per cent alcohol, and immediately pour it off and add 

 absolute alcohol. The 95 per cent alcohol should not act for more 

 than 5 or 10 seconds, its only function being to save the more expensive 

 absolute alcohol. From 10-30 seconds will usually be long enough for 



