In Vivo Imaging of Membrane-Associated Glycans in Developing Zebrafish
Scott T. Laughlin, Jeremy M. Baskin, Sharon L. Amacher, Carolyn R. Bertozzi

Supporting Online Material

This supplement contains:
Materials and Methods
SOM Text
Figs. S1 to S14
References

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Other Supporting Online Material for this manuscript includes the following:
(available at www.sciencemag.org/cgi/content/full/320/5876/664/DC1)
Movies S1 to S9

Movie s1
Z-series of two-color labeling of the mouth region (frontal view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted, starting at 60 hpf, with DIFO-647 and, subsequently, DIFO- 488 using the 2-h time resolution protocol described in Fig. 3A–D. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, brightfield, fluorescence merge (red: DIFO-647; green: DIFO-488). Scale bar: 10 μm.

Movie s2
Z-series of two-color labeling of the head and jaw region (lateral view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted, starting at 60 hpf, with DIFO-647 and, subsequently, DIFO- 488 using the 2-h time resolution protocol described in Fig. 3A–D. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, brightfield, fluorescence merge (red: DIFO-647; green: DIFO-488). Scale bar: 10 μm.

Movie s3
Z-series of two-color labeling of the head and jaw region (ventral view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted, starting at 60 hpf, with DIFO-647 and, subsequently, DIFO- 488 using the 3-h time resolution protocol described in Fig. 3E–H. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, brightfield, fluorescence merge (red: DIFO-647; green: DIFO-488). Scale bar: 100 μm.

Movie s4
Zoom-in of z-series of two-color labeling of the head and jaw region (ventral view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted, starting at 60 hpf, with DIFO-647 and, subsequently, DIFO-488 using the 3-h time resolution protocol described in Fig. 3E– H. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, brightfield, fluorescence merge (red: DIFO-647; green: DIFO-488). Scale bar: 25 μm.

Movie s5
Z-series of two-color labeling of the olfactory organ (frontal view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted, starting at 60 hpf, with DIFO-647 and, subsequently, DIFO- 488 using the 3-h time resolution protocol described in Fig. 3E–H. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, brightfield, fluorescence merge (red: DIFO-647; green: DIFO-488). Scale bar: 10 μm.

Movie s6
Z-series of three-color labeling of the head and jaw region (lateral view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted sequentially, starting at 60 hpf, with DIFO-647, DIFO-488, and DIFO-555 using the 12-h time resolution protocol described in Fig. 3I–N. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, DIFO-555 fluorescence brightfield, fluorescence merge (blue: DIFO-647; green: DIFO-488; red: DIFO-555). Scale bar: 100 μm.

Movie s7
Z-series of three-color labeling of the head and jaw region (ventral view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted sequentially, starting at 60 hpf, with DIFO-647, DIFO-488, and DIFO-555 using the 12-h time resolution protocol described in Fig. 3I–N. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, DIFO-555 fluorescence brightfield, fluorescence merge (blue: DIFO-647; green: DIFO-488; red: DIFO-555). Scale bar: 100 μm.

Movie s8
Z-series of three-color labeling of a hair cell (frontal view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted sequentially, starting at 60 hpf, with DIFO-647, DIFO-488, and DIFO-555 using the 12-h time resolution protocol described in Fig. 3I–N. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, DIFO-555 fluorescence brightfield, fluorescence merge (blue: DIFO-647; green: DIFO-488; red: DIFO-555). Scale bar: 2.5 μm.

Movie s9
Z-series of three-color labeling of the olfactory organ (frontal view). Zebrafish embryos were metabolically labeled with Ac₄GalNAz using the general procedure and reacted sequentially, starting at 60 hpf, with DIFO-647, DIFO-488, and DIFO-555 using the 12-h time resolution protocol described in Fig. 3I–N. Panels (from left): DIFO-647 fluorescence, DIFO-488 fluorescence, DIFO-555 fluorescence brightfield, fluorescence merge (blue: DIFO-647; green: DIFO-488; red: DIFO-555). Scale bar: 10 μm.

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