454 ON THE ORIGIN AND EARLY DEVELOPMENT 



instances the sections were cut parallel to the surface. By this means the greater 

 part of the lymphatic plexus was contained in a few sections and parts could be 

 reconstructed which would be completely lost in cross-sections. The sections were 

 cut 10 and 15 n in thickness, stained on the slide with Ehrlich's hematoxylin, and 

 counterstained with a mixture of eosin, orange G, and aurantia. By this staining 

 method the nucleoli of the lymphatic endothelial cells are reddish in color, which 

 is an aid in distinguishing them from the mesenchyme cells whose nucleoli are 

 bluish-purple. This same contrast was obtained by the use of Mann's methyl-blue 

 eosin stain. 



Such sections were studied by means of oil-immersion reconstruction. The 

 blood-vessels could be identified by the presence of India ink, and the other vessels 

 were all carefully drawn, every strand of endothelium, every nucleus and nucleolus 

 being recorded. The structures were not studied in this manner until extensive 

 investigations of the lymphatics and blood-vessels of this region had been carried 

 out in living and injected embryos. Familiarity with the region as a preparation, 

 and the modification of parallel sections, the constant use of the oil-immersion 

 lens, and a criterion for distinguishing endothelial cells enabled us to detect definite 

 vessels and even a plexus at stages in which our earlier studies and those of other 

 investigators gave no hint of the existence of lymphatics. Needless to say this 

 method is extremely tedious. A drawing of all the endothelial cells, including 

 nuclei and nucleoli, was made of each section, and the drawings of successive sec- 

 tions were combined in graphic paper reconstructions. 



LYMPHATICS OF POSTERIOR LYMPH-HEART REGION IN CHICKS OF 5 TO 6 DAYS 



As has been stated, the pulsation of the posterior lymph-heart can be seen in 

 chicks of 6 to 6| days, a stage when injection shows that the heart is still in the 

 form of a plexus. This lymph-heart plexus connects medially with the first five 

 intersegmental coccygeal veins, and ventrally with a superficial lymphatic plexus 

 which spreads out over the pelvis and anterior body- wall, and is continuous from 

 the tail to the axilla. Cleared, injected specimens show that this superficial plexus 

 has connections through the axillary region with the deep lymphatic plexus located 

 dorsal to the anterior and posterior cardinal veins, near their junction at the duct of 

 Cuvier, with which veins it communicates at a number of places. (A drawing of 

 this plexus has been published in an earlier article E. L. Clark, 1912.) In a chick 

 of 5 days and 18 hours this continuous superficial plexus can be injected and most 

 of it can be seen in the living chick because of the stagnant blood present in its 

 lumen. The lymph-heart has not started to beat at this stage. 



In younger embryos (5 to 5| days) blood-filled lymphatics are also visible, but 

 a continuous plexus over the surface of the body can not be seen in the living or 

 injected specimens. Instead, a plexus is found anteriorly, near the region of the 

 thoraco-epi gastric vein, and connected through the axillary region with the deep 

 jugular plexus and with the veins, as in the older specimen; also another posterior 

 plexus connected with the intersegmental coccygeal veins and confined to the 

 region of the posterior lymph-heart and to the neighboring area over the posterior 



