148 Victor E. Emmel. 
sharply contrasted with any surrounding tissue. Using this stain 
reaction as an index of differentiation, serial sections were studied 
and graphic reconstructions made of different stages in the regenera- 
tion of the limb (Figs. 31-36). 
a. Two Days and Twenty-two Hours after Operation (fig. 31).— 
Although externally at this time there was little, if any, indication 
of segmentation, internally there was found a well defined invagina- 
tion of epidermal cells, within which a differentiation of chitin (7) 
was already evident. This invagination marks the beginning of the 
first and second distal segments. It is here that the chitin plate is 
formed for the flexor muscle of the dactyl, which, it is to be observed, 
is normally not only much larger than the opposing extensor, but 
is also the largest muscle of the limb.* 
b. Four Days and Sia Hours (ig. 32).—The differentiation of 
additional chitin plates is not evident until on the fourth day of 
regeneration. At this time two more invaginations were beginning 
to develop chitin. Of these two invaginations the one representing 
the extensor for the dactyl (e') showed as yet but a slight differen- 
tiation of this tissue. The other invagination (7°) represents the 
flexor for the meropodite, and shows a well-defined plate of chitin. 
In regard to the latter invagination (7?) there are two important 
points to be emphasized. - First, the invagination is not in the next 
or third proximal segment, but in the fourth segment or meropodite ; 
and second, of the two muscles in the meropodite, the invagination 
represents the flexor.® 
‘This invagination occurs at one side of the apex of the bud, so that at 
first the dactyl is relatively Jarger than the index,—a relation, it is interesting 
to note, which corresponds with the condition found in the larval stages of 
development, but is reversed in the adult. (Hmmel, ’06*). 
°In the external segmentation of the limb at this stage, the groove between 
the second and third segments is quite as well marked, if not more so, than 
the groove between the third and fourth segments. This agrees with Haseman’s 
(07) observations on the cheliped, and with my own earlier description (’06) 
for the lobster, that the external segmentation appears to proceed in a disto- 
proximal direction; but the crucial fact that in the present case it is the 
tendon in the fourth segment instead of in the third segment which differ- 
entiates next after the second segment, demonstrates that the external seg- 
mentation cannot, therefore, be safely relied upon alone as a correct index 
of internal differentiation. 
