Fig. S5.

Confirmation of vimentin on the cell surface of senescent cells using commercial antivimentin antibody. (A) SACE analysis using indicated antibodies to detect immunoreactivity toward live senescent NDF cells. An HRP-conjugated IgG or IgM specific secondary antibody was used in conjunction with TMB substrate for colorimetric ELISA reading at an absorbance of 450 nm to detect binding antibodies to the surface of cells. DNA was stained with cell permeable Hoechst to normalize TMB absorbance values. Values (arbitrary units) and SDs were calculated from triplicates. An unpaired Student’s t test was used for statistical analysis. (B) SACE analysis (as in A) using indicated antibodies to detect immunoreactivity toward methanol fixed and permeabilized NDF cells (untreated or IR-treated). An unpaired Student’s t test was used for statistical analysis. (C) ELISA measuring fold increase in vimentin and FSP in membrane lysates from untreated and IR-treated NDFs. (D) Immunostaining of live untreated and bleomycin-treated mLFs stained in suspension with IgM 9H4 clone (red). DNA was stained with DAPI (blue). Arrows highlight 9H4 accumulation at the plasma membrane in bleomycin-treated cells. (Scale bar, 5 μm.)