As an extension of previous investigations, the possibility of the yolk spheres of chick embryo developing into cells under normal and experimental conditions was studied in further detail. It was observed that the deeper cells of the early blastoderm made their morphological differentiation and gra- dually became separated by delamination from the superficial cells to form the primary entoderm; the yolk spheres that entered the subgerminal cavity, although located adjacent to or even intermingled with the deeper cells of the blastoderm, became disintegrated in later stages and did not take any part in the process of entoderm formation. The so-called "columnar epithelial cells" of the germ wall and of the yolk sac lining were not derived from the cells or the yolk spheres of that region, but were transformed from a structure of multinucleated syncytium, with the yolk sphere or granules in its meshes, which is histologically characteristic of the germ wall and the area vitellina interna of the yolk sac. They had an uncertain number of nuclei (from 1 to 3, or none at all) and an extraordinary large body size incomparable to any kind of embryonic cells (average 40-50? in length and 10-15? in diameter; the larger ones even reach 75-80? in length and 18? in diameter). The formation of these "cells" was found to be parallel with the expansion of the vitelline blood vessels along the yolk sac wall. Local cauterization indicated that outside the burned region in which the blood vessels were absent, the "columnar cells" failed to differentiate subsequently. The arrival of the blood vessels seems to induce the surrounding syncytial structure and yolk spheres to arrange themselves from an irregular state into long columns, and in such a way that the syncytial protoplasm and nuclei were moulded by the yolk spheres into a columnar structure, the so-called "columnar epithelial cells", but actually only a structure of syncytium. During this process, the yolk spheres underwent disintegration and liquefaction; some of them broke down into granules, droplets, and fragments of unequal sizes, with positive Feulgen's nuclear reaction. Some even showed an appearance of a pyknotic nucleus; but judging from their morphological detail and disintegration in the sucessive stages, they did not really show the characteristics of a nucleus. These results furnished further evidence that the yolk spheres of chick embryo could not develop into any kind of embryonic cell.

Saline solution (0.9%),containing 5—10?c of radiosulphate(S~(35))——methio-nine(0.2—0.4ml)was injected into the yolk sacs of unincubated to 8—day chickembryos.Fixation was made after 6—24 hours of incubation in various fixa-tives (Bouin's,Carnoy's,neutral formalin,methyl or 95% ethyl alcohol).Mostof the embryos were stained in toto with Ehrlich's hematoxylin and eosin,butsome were stained with Wallback's Giemsa mixture of Mallory's modificationafter sectioning.Serial sections(5—7?) were used for contrast autoradiography.It was observed that a remarkable radiosulphate uptake in the embryonic tissuefollows the cellular pattern and was concentrated in proportion to the cellulardensity of tissue,the tracer is noticeable in the autographs by its ?-tracts andgrains over cytoplasm and nuclei of cells;while the yolk spheres entered thesubgerminal cavity and in the space between ectoderm and entoderm in thegerm wall were not radioactive.The blood vessels,erythroblasts and the mul-tinucleated syncytium,including the so-called “columnar epithelial cells” of theyolk sac lining and of the germ wall,also show high incorporation of radio-sulphate,but not at all in the yolk material.Furthermore,the yolk granules,droplets of unequal size with positive Feulgen reaction,and those even withan appearence of pyknotic nucleus in the “transitional zone” of yolk sac liningas described in the previous investigation,also show no tracer uptake.It isobvious,that radiosulphate is not incorporated into the yolk material.Fromthe above,it may be assumed that no protein synthesis or resynthesis(“??-????????”) occurs in the yolk sphere.These results are in good agreementwith our previous findings and afford further evidence to the conclusion that theyolk spheres are not a living substance incapable of developing into embryoniccell.