Upregulation of PTP-RO protein expression in megakaryocytic cell lines. (A) Total cell lysates containing equal amounts of protein (100 μg) from CTS, CMK, Dami, and Mo7e cells were separated on 7.5% SDS-PAGE. Immunoblotting was performed using anti–PTP-RO antibodies (R5702; 7 μg/mL) or control preimmune antibody (7 μg/mL). (B and C) CMS, CMK, and Dami cells were cultured in RPMI-1640 or DMEM medium (referred to in Materials and Methods) containing 10% FCS in the presence of 100 nmol/L PMA at a concentration of 106 cells/mL. After culture for 0, 24, and 48 hours, cells were collected and lysed with the RIPA buffer. One hundred micrograms of protein from clarified cell lysates were analyzed by 4% to 12% SDS-PAGE followed by Western blotting using two anti–PTP-RO antibodies R5702 (7 μg/mL; B) and D2712 (8 μg/mL; C) as well as control preimmune antibodies (8 μg/mL). The positions of migration of full-length PTP-RO and putative proteolytic PTP-RO are indicated.
