Cys shotgun labeling of macrophages adhering to and engulfing Ig-opsonized cells

A method is being developed to study cytoskeletal reorganization in cell adhesion processes. The initial model process is adhesion and phagocytosis of beads or red blood cells by macrophages. Live cell labeling with Cys reactive fluorophores is performed before and during phagocytosis with different color labeling dyes. Since Cys is a relatively hydrophobic amino acid, its differential exposure and labeling in principle reflects changes in tertiary or quaternary structure of specific proteins. Similar studies conducted on red blood cells under fluid shear conditions showed that specific domains in spectrin undergo extensible unfolding within sheared cells. The initial work here with macrophages also suggests some structural changes in phagocytosis although the proteins and specific sites have yet to be identified.