FIGURE 1.

Establishment of K562 cell lines stably expressing P2X7. K562 leukemia cells were transfected with a blank, wild-type P2X7-expressing, or N187D P2X7-expressing vector. Stably transfected polyclones (K562-V, K562-W, and K562-M) were obtained by G418 selection. A, upper, their successful establishment were confirmed by RT-PCR using primers for either P2X7 or the neo gene. GAPDH was used as an internal control. Lower, the expression of P2X7 in the cells was detected by Western blotting as described under “Experimental Procedures.” β-Actin was used as an internal control. B, flow cytometry analysis of membrane P2X7 levels was performed using FITC-labeled polyclonal antibodies against the extracellular part of P2X7. C, confocal microscopy analysis of P2X7 expression was performed using FITC-labeled polyclonal antibodies against the C-terminal part of P2X7.