부산대학교 도서관

Monoclonal antibodies were raised against a fusion protein consisting of a fragment of 141 amino acids of the C-terminal region of the rat brain voltage-gated K+-channel protein (RCK1) and the λN protein (fusion protein I). Selection of K+-channel-specific hybridoma cell lines was per- formed by means of an ELISA emPloying a fusion protein consisting of the K+-channel-specific peptide sequence and glutathione S-transferase (fusion protein II). For final selection of RCK1 isoform-specific antibodies, a panel of Xenopus oocytes was employed, each injected with cRNA coding for a specific RCK isoform (RCK 1, 2, 4 or 5). Several days after injection, cryosecfions of embedded oocytes were obtained and were employed in immunohistochemical analysis of antibody binding. Of five hybridoma supernatants from stable growing hybridoma cell lines, selected by the fusion-protein ELISA, one monoclonal antibody (denoted K1 C3) recognized exclusively the RCKI- protein isoform, with the other four exhibiting different levels of cross-reactivity with other K‾- channel isoforms, or with unknown protein(s) of non-injected oocytes. The expression of the RCK1 protein in the postnatal brain was studied using, as far as we are aware, the first example of the application of such isoform-specific antibodies. [ABSTRACT FROM AUTHOR]