Immunoblotting of Kell protein in RBCs and transfectants. (A) RBC membrane preparations were separated on a 4-12% gradient SDS-PAGE under reducing conditions and immunoblotted with a polyclonal anti-Kell (see Materials and Methods). Lane 1, Kell null RBCs; lane 2, Kp(b+) (wt) RBCs; lane 3, Kp(a+) RBCs. (B) Protein extracts from 293T transfected cells and RBC membrane preparations were separated on SDS-PAGE under reducing conditions and immunoblotted with the polyclonal anti-Kell. Lane 1, mock-transfected cells; lane 2, Kp(a+) RBCs; lane 3, Kp(b+) (wt) RBCs; lane 4, cells transfected with pRc/CMV-wt Kell; lane 5, cells transfected with pRc/CMV-Kp^a; lane 6, cells transfected with pRc/CMV-Kp^c. The amount of protein loaded in lanes 1, 2, and 3 in (A) was threefold more than in (B) lanes 2 and 3.