FIG 2.

(A) Detection of vimentin expression on the cell surface of U251, RD, Vero, and HeLa cells by flow cytometry. Cells were fixed and incubated with either mouse IgG (black line) or antibody to vimentin (red line). The cells were then incubated with the fluorescent secondary antibody and subjected to flow cytometry analysis as described in Materials and Methods. y axis (counts) = cell counts; x axis (FL1-H) = fluorescence density. (B) Analysis of the cell surface distribution of vimentin and cell surface-bound EV71 by indirect immunofluorescence in U251 cells. Cells were infected with EV71 BrCr (+EV71) at 4°C for 1 h and then stained with specific antibody to either EV71 (red) or vimentin (green) and analyzed by confocal fluorescence microscopy. Bar = 20 μm. (C) Analysis of the distribution of cell vimentin by indirect immunofluorescence in U251 cells (−EV71). Cells were fixed and permeabilized as described in Materials and Methods. Cells were then stained with antibodies to vimentin (Vim; green fluorescence) and EV71 (red florescence) and subjected to confocal microscopy analysis. An overlay of the vimentin and EV71 florescence is also shown (merged). Cell morphology (phase) was assessed by light microscopy.