PRODUCTION AND CHARACTERIZATION OF MULTIPLE ANTIGENIC PEPTIDE ANTIBODIES TO THE ADENOSINE A(2B) RECEPTOR

A polyclonal antibody to the human adenosine A(2b) receptor (A(2b)R) w as produced by immunizing a chicken with a multiple antigenic peptide consisting of eight copies of a 16-amino acid peptide, corresponding t o the presumed second extracellular loop of the A(2b)R, linked to a br anched lysine core. Western blotting with affinity-purified antibody r evealed the human A(2b)R to be a protein of approximately 50-55 kDa, f ound in a variety of tissues including thymus, colon, and small intest ine. The antibody also recognized mouse and rat A(2b)Rs and revealed h eterogeneity in size, with a 35-kDa protein being detected in small in testine in addition to the larger 50-52-kDa species in thymus, colon, and placenta. The chicken anti-human A(2b)R peptide antibody recognize d the receptor in both frozen and formalin-fixed tissue sections. In h uman colon, the A(2b)R was highly expressed in epithelial cells of the crypts. A(2b)R immunoreactivity was also apparent in syncytiotrophobl ast cells of human placental villi and in the basal zone of murine cho rioallantoic placenta. These cell type-specific patterns of expression are consistent with the hypothesized roles of the A(2b)R in mediating electrogenic Cl- secretion and the resulting secretory diarrhea cause d by colonic crypt abscesses and in regulating morphogenesis of the pl acenta. Insight into the multiple physiological consequences of A(2b)R engagement will be forthcoming from an analysis of the cell type-spec ific expression of this receptor in additional tissues.