Hematopoietic specification and specific cell lineages in the zebrafish embryo. (A-E) Markers of hematopoietic specification. (A) The earliest site of primitive hematopoiesis can be identified in the posterior lateral plate mesoderm using a riboprobe for tal1(scl) (blue, open arrowhead). (B) gata1 expression (blue) in the developing intermediate cell mass (ICM). (C) spi1 (pu.1) expression (blue) in the anterior lateral plate mesoderm (open arrowhead), below the head (closed arrowhead), defines the region producing the first wave of primitive macrophages. (D,E) Definitive hematopoiesis, marked by cmyb and runx1 (blue), commences in the ventral wall of the dorsal aorta (arrowheads). (F-K) Markers of specific hematopoietic cell lineages. (F) Lineage-committed erythroid cells expressing hbbe3 (globinβe₃) (blue) in the ICM and circulating cells over the yolk. (G) mpx-expressing granulocytes (blue) in circulation over the yolk and in the ventral tail (open arrowheads). (H) Sudan black cytochemistry marks the same dispersed granulocyte population (open arrowheads). (I) ighm (IgM) expression in a B lymphocyte (open arrowhead) in a kidney section, adjacent to a renal tubule (closed arrowhead). (J) T lymphocytes marked by rag1 expression (blue) in the bilateral thymi. (K) CD41 (itga2b) expression (blue) in circulating thrombocytes over the yolk (open arrowhead). (A-G,I-K) Gene expression by whole mount in situ hybridization. Microscopy was performed using a Nikon SMZ-1500 microscope (Nikon, Melville, NY) equipped with a 0.75-11.25× objective (A-H,J-K) and a Nikon Optiphot-2 microscope equipped with a 100×/1.40 oil objective (I). Images were obtained using a Zeiss Axiocam MRc5 digital camera (Carl Zeiss, Thornwood, NY) with Axiovision AC software (Release 4.5). Images were processed using Adobe Photoshop CS2 9.0 and Adobe Illustrator CS2 12.0.1 (Adobe Systems, San Jose, CA).