Load-independent ceiling of single-target phagocytic membrane extension revealed by microneedle backtracking assay in macrophages

The zipper model describes the ligand-receptor-driven progression of the phagocytic cup during macrophage engulfment. However, whether the maximum engulfment achievable for a single target is altered by prior or concurrent phagocytic events (i.e., intracellular phagocytic load) remains unclear. Here, we used IgG-coated, nondigestible glass microneedles as standardized Fcγ receptor ligands and defined the single-target engulfment ceiling as the membrane extension length at which backtracking begins. We then tested whether this ceiling changes after macrophages internalize increasing numbers of IgG-coated polystyrene beads. Across cells, the maximum membrane extension on a microneedle was quantitatively unchanged regardless of the number of internalized indigestible beads. Within the same cell, additional bead ingestion - up to the maximal bead-phagocytosis limit - did not measurably alter the maximum extension achieved on a microneedle. These data establish a load-independent ceiling for single-target engulfment. This invariance suggests that local membrane recruitment and extension are regulated independently of the cell-wide phagocytic burden, supporting a spatially compartmentalized control mechanism that decouples single-target membrane extension from the total intracellular cargo load.