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Fig. 4. Expression of vasculogenesis and hematopoiesis markers in Ihh–/– ES cell embryoid bodies. PECAM whole-mount immunocytochemistry in day 13 Ihh+/– embryoid bodies reveals expression in endothelial cells lining the interior of blood islands (A, arrows) and in core patches, whereas Ihh–/– embryoid bodies express PECAM in only core patches. Scale bar: 200 µm. (B) RT-PCR analysis examining expression of vasculogenesis and hematopoiesis markers. Ihh+/– embryoid bodies upregulate the markers Tal1, Flk1 and Flt1 during blood island development (days 10-13), whereas the loss of Ihh results in a dramatic reduction in mRNA levels for these genes. HPRT is shown as a loading control.