Figure 2.
Figure 2. Immunohistochemistry with antiglobin antibodies. (A) E10.5 yolk sac stained with anti–βH1-globin antibody. Positive primitive erythroblasts are evident throughout a large blood vessel, whereas adjacent mesoderm-derived (m) and endoderm (en) cells are negative. The scale bar represents 25 microns. (B-C) E12.5 neighboring embryo sections stained with anti–βH1-globin (B) and anti–βmajor-globin (C) antibodies. The anti–βH1-globin antibodies decorate primitive erythroblasts in blood vessels (white arrows). Anti–βmajor-globin antibodies decorate both primitive red cells in vessels and definitive erythroblasts maturing in the liver (liver). Int indicates intestine. The scale bar represents 50 μm. (D) E13.5 peripheral blood cells stained both with anti–βH1-globin antibodies and with DAPI (left). A paired Hoffman modulation contrast image (HMC, right) shows the βH1-globin–positive nucleated cells (black arrows), βH1-globin–negative enucleated cells (blue arrows), and βH1-globin–positive enucleated cells (red arrows). The scale bar represents 10 μm. (E) E15.5 peripheral blood cells stained with anti–βH1-globin antibodies (left). A paired Hoffman modulation contrast image (HMC, right) of the same field of cells indicates a βH1-globin–positive nucleated cell (black arrow), the βH1-globin–negative enucleated cells (blue arrows), and a βH1-globin–positive enucleated cell (red arrow). The scale bar represents 25 μm. (F) Morphometric analysis of cell area of 3 populations of peripheral blood cells at E15.5 of mouse gestation. Nucleated (black square) and enucleated (red diamond) βH1-globin–positive cells are both similar in size and are significantly larger than βH1-globin–negative definitive erythrocytes (blue circle). Ordinate units are μm2. (G) Immunohistochemical analysis of βH1-globin expression in circulating blood cells from E11.5, E14.5, and E17.5 mouse embryos (right column) with paired phase contrast images (left column). At E14.5, both nucleated and enucleated (arrow) βH1-globin–positive cells are evident. All images are at the same magnification. Scale bar represents 10 μm.

Immunohistochemistry with antiglobin antibodies. (A) E10.5 yolk sac stained with anti–βH1-globin antibody. Positive primitive erythroblasts are evident throughout a large blood vessel, whereas adjacent mesoderm-derived (m) and endoderm (en) cells are negative. The scale bar represents 25 microns. (B-C) E12.5 neighboring embryo sections stained with anti–βH1-globin (B) and anti–βmajor-globin (C) antibodies. The anti–βH1-globin antibodies decorate primitive erythroblasts in blood vessels (white arrows). Anti–βmajor-globin antibodies decorate both primitive red cells in vessels and definitive erythroblasts maturing in the liver (liver). Int indicates intestine. The scale bar represents 50 μm. (D) E13.5 peripheral blood cells stained both with anti–βH1-globin antibodies and with DAPI (left). A paired Hoffman modulation contrast image (HMC, right) shows the βH1-globin–positive nucleated cells (black arrows), βH1-globin–negative enucleated cells (blue arrows), and βH1-globin–positive enucleated cells (red arrows). The scale bar represents 10 μm. (E) E15.5 peripheral blood cells stained with anti–βH1-globin antibodies (left). A paired Hoffman modulation contrast image (HMC, right) of the same field of cells indicates a βH1-globin–positive nucleated cell (black arrow), the βH1-globin–negative enucleated cells (blue arrows), and a βH1-globin–positive enucleated cell (red arrow). The scale bar represents 25 μm. (F) Morphometric analysis of cell area of 3 populations of peripheral blood cells at E15.5 of mouse gestation. Nucleated (black square) and enucleated (red diamond) βH1-globin–positive cells are both similar in size and are significantly larger than βH1-globin–negative definitive erythrocytes (blue circle). Ordinate units are μm2. (G) Immunohistochemical analysis of βH1-globin expression in circulating blood cells from E11.5, E14.5, and E17.5 mouse embryos (right column) with paired phase contrast images (left column). At E14.5, both nucleated and enucleated (arrow) βH1-globin–positive cells are evident. All images are at the same magnification. Scale bar represents 10 μm.

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