Monoclonal antibodies distinctively recognizing the subtypes of inositol 1,4,5-trisphosphate receptor: Application to the studies on inflammatory cells

AbstractMonoclonal antibodies were raised that specifically recognize the COOH-terminal sequences and the loop sequences between the fifth and the sixth transmembrane spanning regions of human inositol 1,4,5-trisphosphate receptor (IP3R) type 1, 2 and 3. Western blot analysis using Jurkat cells, mouse cerebellum, COS-7 expressing IP3R type 3 cDNA showed that those monoclonal antibodies reacted specifically with each of these three IP3R subtypes and that they do not cross-react. These antibodies could be used for the specific immunoprecipitation of IP3Rs. Using these monoclonal antibodies, the expression profiles of IP3R-subtype proteins were found to be different among inflammatory cells such as macrophages, polymorphonuclear cells, mast cells, eosinophils, splenocytes, thymocytes and megakaryocytic cells. Usually, more than one type of IP3R were expressed in a cell simultaneously. The observation of CMK cells under immunofluorescence confocal microscopy revealed that IP3R type 1 and type 2 are located at different subcellular fractions