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. 2014 Apr 11;24(7):619–637. doi: 10.1093/glycob/cwu027

Fig. 1.

Fig. 1.

Recombinantly engineered Lec8 cells express surface-bound LDN and LDNF. (A–C) Lec8 (white bars), Lec8-GalNAcT (L8-GT; gray bars) and Lec8-GalNAcT-FucT (L8-GTFT; black bars) were fixed and stained with biotinylated lectins (A) and mouse monoclonal antibodies (B and C) for flow cytometry analysis. (A) Data are expressed as fold change in geoMFI over the negative control, SA-488, represented on the graph as a dotted line. Mean ± SEM of two independent experiments are shown and two-way ANOVA with Tukey's multiple comparisons test was used to determine significance. (B) A representative shift with each monoclonal antibody is shown. (C) Cells were treated with (hashed bars) or without kifunensine for 8 days to disrupt complex N-glycan synthesis before fixation. Data are expressed as fold change in geoMFI over the negative control—mouse IgG1 isotype control for (B), and IgG1 isotype control or normal mouse IgG for (C). Error bars represent SEM of two (C—left) or three (C—right) independent experiments and one-way ANOVA with Tukey's test were used. (D) SDS–PAGE and western blotting were performed on cell and Schistosoma mansoni life stage lysates and using the monoclonal antibodies, with detection by HRP-conjugated secondary antibodies and chemiluminescent substrate on film. Primary antibody concentrations and exposure times are specified for each pane. The six-point star denotes bands that are not antibody-specific, since they appear with secondary antibody and substrate only when SEA and adult worm lysates are stained, likely the result of trace amounts of mouse immunoglobulins in the parasite preparations. SA, streptavidin; ConA, concanavalin A; GSL-II, Griffonia simplicifolia lectin II; WFA, Wisteria floribunda agglutinin; AAL, Aleuria aurantia lectin; Kif, kifunensine; Iso, isotype control antibody or normal mouse IgG; stds, molecular weight standards; Cerc., cercarial lysate; Som., schistosomula lysate; Adult, adult worm lysate; SEA, soluble egg antigen; ns P ≥ 0.05; *P = 0.01–0.05; **P = 0.001–0.01; ***P = 0.0001–0.001; ****P < 0.0001.