Fluorescence imaged microdeformation (FIMD) analysis of RhAG on a normal erythrocyte.

(A) Image of a micropipette-aspirated erythrocyte with fluorescently labeled RhAG. The schematic illustrates an aspirated cell of projection length L in a pipette of radius R_p. The entrance, e, and cap, c, regions are identified. (B) Relative fluorescence intensities or densities averaged across the entrance and cap for 9 cell projections of various lengths. See “Materials and methods” for normalization method. (C) Ratio of entrance-to-cap densities as a function of projection length. This plot (with linear fit forced to interceptρ_e/ρ_c = 1) illustrates the increasing component gradients that are characteristic of considerable cytoskeletal attachment.21 For typical bilayer probes, the slope, m, has a value close to 0.0, whereas freely diffusing proteins such as GPI-linked CD59 give definitively negative slopes.