Fig. 3. Distribution of endothelial cells in Hand1 mutant yolk sacs. Hand1 is expressed in the mesodermal layer of the yolk sac but not in differentiated endothelial cells. (A-F) Co-localization of PECAM protein and Hand1 expression. (G,H) Phase contrast images of A-F. Immunohistochemistry was performed with yolk sac sections of 8.5 dpc Hand1^ß-gal/+ (A,C,E,G) and 9.5 dpc Hand1^{ß-gal/ß-gal} (B,D,F,H). Anti-ß-gal antibody was used to visualize HAND1-expressing cells (A,B). Both yolk sacs expressed PECAM (C,D). end, extra-embryonic endoderm; mes, extra-embryonic mesoderm. Asterisks in Gindicate blood vessels. (I,J) Visualization of tissues expressing PECAM in yolk sacs. Whole-mount staining of 8.5 dpc Hand1^+/+ (I), 9.5 dpc Hand1^{ß-gal/ß-gal} (J) and 9.5 dpc Hand1^+/+ littermate (K) yolk sacs with anti-PECAM antibody. Wild-type yolk sacs show integration into large vessels in both 8.5 dpc and 9.5 dpc (arrowheads). Hand1^{ß-gal/ß-gal} yolk sac lacks integrated vasculature.