Extended Data Fig. 8 |. Siglec-7 and -9 undergo rapid internalization induced by (SigL^bio)₄-SA tetramer and restore quickly on cell surface.

a,b, U937-derived Siglec-7⁺ (a) and -9⁺ (b) macrophages were incubated with (8^bio)₄-SA-AF488 at 37 °C over the indicated time periods, then stained with anti-Siglec-7 and anti-Siglec-9 antibodies, followed by flow cytometry analysis. c,d, Fluorescence microscopy imaging showing that the internalized Siglec-7 (c)/Siglec-9 (d) and AF488 dye were mostly colocalized with the early endosome marker (Rab5). Scale bar, 10 μm. e,f, Fluorescence microscopy imaging illustrating the internalized Siglec molecules and AF488 dye in U937-derived macrophages disappear quickly following the wash away of (8^bio)₄-SA tetramer. Scale bar, 10 μm. g,h, Quantification of the rapid restoration of cell surface Siglec-7 (g) and -9 (h) levels on U937-derived macrophages after the removal of (8^bio)₄-SA tetramer. Data are mean ± s.d. Two-tailed unpaired Student’s t-test. ns, not significant (g,h).