Production of antisera to synthetic decapeptide of the Cooh-Terminus of rat glut2

Abstract

In the present study, the production of an anti-GLUT 2 antibody is reported. The antibody (S-5096) was raised in New Zealand rabbits immunized with an immunogen prepared by cross-linking the synthetic peptide and a carrier protein. A COOH-terminal decapeptide of rat GLUT 2 predicted sequence was synthesized and coupled to keyhole limpet hemocyanin, using the glutaraldehyde method. Subcellular membrane fractions were prepared from renal cortex (C) and medulla (M), and subjected to Western blotting analysis using the antiserum in a 1:200 dilution and, subsequently, [125I]protein A. As the results showed, the S-5096 anti-serum showed clear blots in kidney samples, stronger in cortex than in medulla as expected, whereas white adipose tissue and heart samples did not show any immunoreactivity. In addition, immunoblots were detected in samples prepared from duodenum and jejunum, as well as from isolated pancreatic B cells. In conclusion, the results clearly show that an anti-Glut 2 antibody, efficient enough to detect the rat protein by Western blotting analysis, was obtained