STAINS AND STAINING 
47 
12. Extract the stain in 1 per cent iron-alum, watching the process under the 
microscope. 
13. Wash several hours in-water. 
14. Alcohol series: 10, 30, 50, 70, 80, 95, 100 per cent. 
If a counter-stain is desired, introduce it in one of the alcohols of 
this series. 
15. Absolute alcohol and xylol, equal parts. 
16. Xylol. 
17. Mount in balsam. 
While these two schedules would enable the student to apply 
the method in case of objects to be mounted whole, like filamentous 
algae, fern prothallia, etc., a complete schedule is given in chapter 
viii on “The Venetian Turpentine Method.” 
The times given above must not be accepted as final. Many 
prefer to wash in water for several hours after the first immersion in 
iron-alum. Some think that 4 hours is enough for the entire process. 
Many put the slide into iron-alum in the morning and finish the pro¬ 
cess in the afternoon. These short schedules are not likely to prove 
satisfactory with mitotic figures. A plan which has proved con¬ 
venient and very successful is to put the slide into the iron-alum 
in the morning, wash in water for an hour at some convenient 
time in the afternoon, leave it in the § per cent haematoxylin over 
night and finish the preparation the next morning. It is a long 
process, requiring care, patience, and judgment, but it is worth the 
effort. 
Chromosomes, centrosomes, and pyrenoids take a brilliant black; 
or, if the second treatment with iron-alum be more prolonged, 
a blue black or purple. Achromatic structures stain purple, but 
the stain can be extracted while it is still bright in the chromosomes. 
Lignified, suberized, and cutinized structures stain lightly or not at 
all. Cellulose does not stain so deeply as with Delafield’s haema¬ 
toxylin. Archesporial cells and early stages in sporogenous tissue 
stain gray. Many details which are not so brilliantly colored often 
show good definition. 
If a counter-stain is desired, anything which gives a serviceable 
contrast may be used. In any case, the haematoxylin stain must be 
complete and the washing thorough before the second stain is applied. 
An aqueous stain should be applied just after the final washing in 
water; an alcoholic stain should be applied during the process of 
