528 Wisconsin Academy of Sciences, Arts, and Letters. 
specially differentiated outer layer, but that be found such a 
layer in a few cases, in starch from the tuber of a potato. 
Sfich a layer is described by Salter (6) who believed that it is 
composed of starch, but that it is much more dense than the 
remainder of the grain, hence its different staining qualities. 
I have taken up the study of this outer differentiated layer 
with special reference to the question of its chemical nature 
and its relation to the growth of the starch grain. 
I have studied material from the rhizome of Ganna, tuber 
of potato', leaves and stems of Pellionia Daveauana, grains of 
com, wheat, barley, rye and sugar-cane, seeds of bean and pea. 
A number of fixatives were tried but Flemming’s weaker and 
stronger chrom-osmic-acetic acid mixture gave the best results. 
The staining was done in all cases where other stains are not 
mentioned, by the safranin, gentian violet, orange and triple 
stain method. 
If one examines sections stained by this method, containing 
hundreds of starch grains large and small, the striking fact is 
noticed that the mass of each grain is a, bright violet, the strata 
being respectively lighter or darker, while 1 around the violet 
mass and inside the leucoplast, a sharply defined orange layer 
is apparent. 
In order to determine in how far the staining reactions were 
due to different lengths of exposure to the individual stains, a 
series of slides was prepared. In all cases they were exposed 
to safranin for five minutes; after washing in water, six slides 
were exposed to a saturated solution of gentian violet for five 
minutes each, then treated with orange for the following differ¬ 
ent lengths of time: one minute, five 1 minutes, ten minutes, 
twenty minutes, sixty minutes, three hours. These prepara¬ 
tions will be referred to by fractions, the numerator represent¬ 
ing the time of exposure to violet in minutes and the denom¬ 
inator the time of exposure to orange. 
In 5/1 the body of the grain is made up of deeply stained 
violet layers. The outer layer is also violet but very pale in 
color. The leucoplasts do not appear on all grains, but when 
visible are grayish in color. 
In 5/5 the body of the grain is again stained dark violet but 
an orange layer at the outside is plainly marked and extends 
entirely around the violet portion, beneath the leucoplast. 
This is the pale violet layer of the 5/1 preparation. 
