194 BOTANICAL GAZETTE [SEPTEMBER 
in running water they were dehydrated through a close series of 
graded alcohols, as follows: 5, 12, 20, 35,50, 70, 85, 95, and 100 
per cent. The xylols were also graded, but less closely: 15, 30, 50, 
70, 85, and roo per cent. The material was infiltrated with 
paraffin by adding the latter a little at a time, and preventing 
actual contact of the paraffin with the material by means of a 
perforated cardboard shelf fitted into the vial, a centimeter above 
the material. 
Longitudinal and cross sections were cut serially 1o uw thick and 
stained by the usual methods employed for iron-alum haema- 
toxylin. A counterstain of gold orange was found very effective 
in bringing out the otherwise transparent walls. The gold orange 
is dissolved in the clove oil to saturation. This is then decanted 
off and about one-fourth the volume of fresh clove oil added. This 
solution is poured on the slide after it has been stained and cleared 
in xylol. Only about a minute is necessary to stain the walls; if 
continued longer it colors the cytoplasm also. The gold orange has 
a great tendency to crystallize out as the oil evaporates, especially 
if the stain is too highly saturated. It is therefore advisable to 
rinse the slide with clove oil, followed by xylol. 
In more recent work it was found that very brilliant prepara- 
tions may be stained with safranin and light green as follows: the 
safranin must be a concentrated solution in 50 per cent alcohol (a 
full strength stock solution was used), with the sections left in it 
1-3 days. After a rapid washing in 50, in 80, and then in 95 per 
cent alcohol the sections were placed in light green (about 1 per 
cent in 95 per cent alcohol) 2-5 minutes. The time for the action 
of the light green varies with the age of the material, the strength of 
the stain, and the length of time the sections were stained in 
safranin. It is desirable, therefore, to stain all the sections of one 
collection at the same time, and not to mix several collections in one 
staining. One or two trials will enable one to determine how long 
to leave the sections in light green, and the remaining slides may be 
carried through by this time schedule. If left in light green too 
long, the safranin will be washed out of the nuclei, and if taken 
out too soon the light green is not impregnated in the cell walls 
sufficiently to give the desired brilliant contrast. From light 
