Glycosylation of c-Mpl WT and mutants. (A) Immunoblot analysis shown of transiently transfected HeLa cells that have been treated 24 hours before harvest with 5 µg/mL Tunicamycin (lanes 2, 5, 8, 11, and 14), 10 µg/mL Tunicamycin (lanes 3, 6, 9, 12, and 15), or without Tunicamycin (lanes 1, 4, 7, 10, and 13), respectively. Untransfected HeLa cells served as controls (lanes 1-3). The c-Mpl was detected using a primary antibody against c-Mpl. Equal loading was controlled by an antibody directed against actin. Separating lines indicate positions in the blot, whereas the order of lanes was changed using Adobe Photoshop Creative Suite 2 for better understanding (original order was: WT, 106, 102, and 104). (B) The analysis of THPO influence on c-Mpl glycosylation is shown. Immunoblot analysis of transiently transfected HeLa cells. Untransfected HeLa cells served as controls. THPO was added 6 hours before harvest at a concentration of 50 mg/mL (lanes 6-10). The c-Mpl was detected using a primary antibody against c-Mpl. Equal loading was controlled by an antibody directed against tubulin.