Figure 5. The glycocalyx of phagocytic targets constitutes a mechanical barrier to ligand recognition and engagement by phagocytic receptors.

A) Diagrammatic representation of experimental model. B) Avidin beads opsonized with anti-avidin antibodies and containing biotin-PEG of indicated sizes were incubated with BMDM for 15 min. Normalized binding index is from >5 fields of 5–10 cells, n=3. Bars represent means ± SEM. C) RAW 264.7 cells and those stably expressing Dectin-1 challenged with heat-killed Candida albicans and stained with calcofluor-white (CFW) and Alexa488-phalloidin (F-actin) after fixation and permeabilization. D) The mean normalized binding index determined for >5 fields of >10 cells each, n=3. E) Diagrammatic representation of C. albicans cell wall. F) Wildtype (wt) and indicated MNN deletion strains of C. albicans live-stained with CFW to detect chitin and concanavalin A (ConA) to detect mannans. G) Quantitation of Alcian blue staining for C. albicans strains shown in F. H) Representative images of RAW 264.7 cells stably expressing Dectin-1 challenged with indicated C. albicans strains stained with calcofluor-white (C. albicans) and Alexa488-phalloidin (F-actin) after fixation and permeabilization. I-J) Quantification of RAW 264.7 cells stably expressing Dectin-1 in I, or parental RAW 264.7 cells in J, and challenged with indicated C. albicans strains. K-L) BMDM challenged with wt and ΔMNN-sextuple C. albicans strains. Representative images are shown in K. Normalized binding index determined for >5 fields of >5 cells, n=3. Bars represent means ± SEM. M) Macrophages elicited to the peritoneal cavity by periodate were challenged with C. albicans strains in vivo for 1 h followed by fixation and staining for F4/80. Normalized binding index determined for >5 fields of >5 cells each, n=3. Bars represent means ± SEM. All scale bars, 10 μm.