Specific detection of non‐functional human P2X7 receptors in HEK293 cells and B‐lymphocytes

Abstract
P2X7 receptor/channels mediate ATP-induced apoptosis in a range of cells including lymphocytes. HEK293 cells were transfected with wild-type human P2X7 receptor or site-directed mutant constructs (K193A, K311A and E496A) known to be non-functional from measurements of barium/ethidium influx in the presence of ATP or 2′,3′-O-(4-benzoylbenzoyl)-ATP. An antibody was designed against an epitope from a loop adjacent to the extracellular ATP site. The epitope was unavailable in cells expressing normal functional surface receptors. Non-functional surface receptors as well as intracellular receptors selectively bound the antibody. So did B-lymphocytes from chronic lymphocytic leukemia patients expressing non-functional (E496A) mutant receptor