50 
METHODS IN PLANT HISTOLOGY 
If, after rinsing in water, the stain is evidently too weak, put 
the slide or section back into the stain until it appears overstained. 
Place the slide in acid alcohol. If an acid alcohol with 2 drops of 
HC1 to 100 c.c. of 70 per cent alcohol reduces the stain too much in 4 
or 5 seconds, use less acid or stain longer. Transfer to 70 per cent 
alcohol without any acid. As soon as the color changes from red to 
purple, examine under the microscope. If it is still overstained, 
return to the acid alcohol; if the stain is too weak, return to the 
haematoxylin and try it again. After the haematoxylin is just right, 
apply a contrast stain, if you wish to double stain. Before trans¬ 
ferring to the xylol wipe the alcohol from the back of the slide, or at 
least rest the corner of the slide upon blotting-paper for two or three 
seconds, in order that you may not carry over so much alcohol into 
the xylol. Add a drop of balsam and a cover. Since the xylol is very 
volatile, this last step must be taken quickly. If blackish spots 
appear they are usually caused by the drying of sections before the 
balsam and cover are added; if there are whitish spots or an emulsion¬ 
like appearance, the clearing is not thorough; this may be caused by 
poor xylol (or other clearing agent); by absolute alcohol which is 
considerably weaker than its name implies (the absolute alcohol 
must test at least as high as 99 per cent, and ought to test as high 
as 99.5 per cent, if xylol is to be used for clearing); or by passing 
too quickly through the absolute alcohol and xylol, or even by 
moisture on the cover-glass. The last danger is easily avoided by 
passing the cover quickly through a Bunsen or alcohol flame before 
laying it on the balsam. 
Delafield’s haematoxylin is the most generally useful stain in the 
haematoxylin group. It brings out cellulose walls very sharply, 
and consequently is a good stain for embryos and the fundamental 
tissue system in general. With safranin it forms a good combination 
for the vascular system, the safranin giving the lignified elements a 
bright red color, while the haematoxylin stains the cellulose a rich 
purple. It is a good stain for chromatin, and the achromatic struc¬ 
tures show up fairly well, but can be brought out much better by 
special methods. Archesporial cells and sporogenous tissue are 
very well defined if proper care be taken. Lignified and suberized 
walls and also starch and chromatophores stain lightly or not at all. 
Whenever you are in doubt as to the selection of a stain for general 
purposes, we should advise the use of Delafield’s haematoxylin. 
