Fig. 1

Internalization of PTD-poMx1 fusion protein. Intracellular localization of PTD-poMx1 fusion protein in ST cell line (a) and 3D4/21 cell line (b) using confocal microscopy. Cells were treated with 80 μg/ml PTD-poMx1 fusion protein for 6, 12 and 24 h at 37 °C, then fixed in 4 % PFA for 15 min at room temperature after three washes with PBS. The anti-poMx1 mAb was applied as a primary antibody for 1 h at 37 °C. After washing three times with PBS, cells were incubated with FITC-goat anti-mouse IgG as a secondary antibody for 30 min at 37 °C. Fluorescent spots attributed to PTD-poMx1 were observed using fluorescence microscopy and recorded with a digital camera. Bar, 10 μm. c The transduction kinetics of PTD-poMx1. ST or 3D4/21 cells were treated with 80 μg/ml PTD-poMx1 and sampled every hour for six hours. Each sample was washed three times with PBS and lysed in cold lysis buffer for 10 min. Total cell lysates were separated by SDS-PAGE, transferred to nitrocellulose membrane, and reacted with antibodies to PTD-poMx1. The blot was stripped and re-probed with anti-β-actin antibody as a loading control. d The stability of PTD-poMx1. ST or 3D4/21 cells were treated with 80 μg/ml PTD-poMx1 for 6 h at 37 °C. After washing three times with PBS, cells in DMEM free from PTD-poMx1 were incubated for an additional 4, 8, 12, 16, and 20 h. Intracellular PTD-poMx1 in the cell lysates was detected using anti-poMx1 mAb as previously above