![Figure 5. In vivo cell labeling with a photoactivatable cell tracer demonstrates the double potential of the primitive (rostral) myeloid progenitors. (A) At 20 hpf, after injection of caged fluorescein-dextran in the embryo at the 1-cell stage, the fluorescein was uncaged (hence its natural fluorescence was restored with a pulsed ultraviolet [365 nm] laser beam) in 10 primitive myeloid progenitors, located along the left lateral border of the pericardium. Fifteen hours later (35 hpf), some of the green-fluorescent labeled cells are observed in the yolk sac circulation valley (). (B) 72 hpf; immunodetection of uncaged fluorescein (AEC staining, red) after SB staining reveals the granulocyte vs. macrophage nature of the progeny of the primitive myeloid progenitors photolabeled in panel A. The drawing recapitulates the locations of all red (uncaged fluorescein-positive) cells identified in this embryo; red dots represent macrophages, stained only by AEC (6/17 cells); these are located along the retina, along the base of the dorsal fin (ii,iii, red arrowheads), in the CHT. Black dots represent cells stained by both AEC and SB (11/17 cells), hence granulocytes; those are located around the eye, ventral to the trunk somites, in the CHT (i,iv,v, black arrowheads). Note that the dextran-coupled uncaged fluorescein revealed by AEC is in the whole cell, whereas SB is confined to the cytoplasm, hence the red label is most apparent in the nucleus. Scale bars, 25 μm. (C) 72 hpf; fluorescent combined immunodetection of uncaged fluorescein (FITC-tyramide, green) with a detection of the peroxidase activity of granulocytes (Cy3-tyramide, red), after uncaging of fluorescein in 10 primitive myeloid progenitors in the yolk sac at 20 hpf. Green dots on the drawing represent macrophages (17/34 cells) that are located in the head mesenchyme, around the eye, in the ganglion cell layer (rgc) of the retina (vi), dorsal to the somites (ii,iii), along the yolk tube (i), and in the CHT (v, vii, green arrowheads). Yellow dots and arrowheads stand for granulocytes, stained in both green and red (17/34 cells). Those cells are located in the head mesenchyme, dorsal to the somites (ii,iii), in the CHT (vii). Scale bars, 50 μm.](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/111/1/10.1182_blood-2007-06-095398/3/zh80020811270005.jpeg?Expires=1724167194&Signature=a9EmgRlSKScpaXbjOBbKVztV-Zsktjdw8MNbi9JCHHKpH4s~tpcujVsKzaTf3a-NlhRlIEy-DSxqsKiPjenmUTBmOucHffSXwPzxFm6cPspvCEMpfJoAWCcz5g7OqSyqAjH4Ko5dUdOkS6p1DOBhv6gqCdNW7z0RuCSDLHaodfA2QvyRYBk~rzv9HPS7CEduRw4sLkNIZgswY8lip-YNmIiHyRfY82zFMRpactMUIKwtxGmj4c3Y2K769CkTqxaYN3i3hOoK7hTePbbo69NbnXV0EM-dIzWJmIOWLQR0jTZdxfd~qXJ3Au5-nUdp5A9JHLuzWafZpUZ8t27R2-9TZA__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
In vivo cell labeling with a photoactivatable cell tracer demonstrates the double potential of the primitive (rostral) myeloid progenitors. (A) At 20 hpf, after injection of caged fluorescein-dextran in the embryo at the 1-cell stage, the fluorescein was uncaged (hence its natural fluorescence was restored with a pulsed ultraviolet [365 nm] laser beam) in 10 primitive myeloid progenitors, located along the left lateral border of the pericardium. Fifteen hours later (35 hpf), some of the green-fluorescent labeled cells are observed in the yolk sac circulation valley (
). (B) 72 hpf; immunodetection of uncaged fluorescein (AEC staining, red) after SB staining reveals the granulocyte vs. macrophage nature of the progeny of the primitive myeloid progenitors photolabeled in panel A. The drawing recapitulates the locations of all red (uncaged fluorescein-positive) cells identified in this embryo; red dots represent macrophages, stained only by AEC (6/17 cells); these are located along the retina, along the base of the dorsal fin (ii,iii, red arrowheads), in the CHT. Black dots represent cells stained by both AEC and SB (11/17 cells), hence granulocytes; those are located around the eye, ventral to the trunk somites, in the CHT (i,iv,v, black arrowheads). Note that the dextran-coupled uncaged fluorescein revealed by AEC is in the whole cell, whereas SB is confined to the cytoplasm, hence the red label is most apparent in the nucleus. Scale bars, 25 μm. (C) 72 hpf; fluorescent combined immunodetection of uncaged fluorescein (FITC-tyramide, green) with a detection of the peroxidase activity of granulocytes (Cy3-tyramide, red), after uncaging of fluorescein in 10 primitive myeloid progenitors in the yolk sac at 20 hpf. Green dots on the drawing represent macrophages (17/34 cells) that are located in the head mesenchyme, around the eye, in the ganglion cell layer (rgc) of the retina (vi), dorsal to the somites (ii,iii), along the yolk tube (i), and in the CHT (v, vii, green arrowheads). Yellow dots and arrowheads stand for granulocytes, stained in both green and red (17/34 cells). Those cells are located in the head mesenchyme, dorsal to the somites (ii,iii), in the CHT (vii). Scale bars, 50 μm.
