Production and characterization of four anti-neuropeptide Y monoclonal antibodies

Human neuropeptide Y (hNPY) is a potent vasoconstrictor peptide of 36 aminoacid residues. We isolated hybridomas secreting four monoclonal antibodies directed against various epitopes of neuropeptide Y and studied their cross-reactivity with peptide YY (PYY) and the pancreatic polypeptide (PP), two peptides sharing sequence homologies with hNPY (respectively 70% and 50%). The antibody NPY02 is an IgGl with a Ka of 5.5 x 1010 liters/mol. It binds to the 11-24 region of NPY (IC50 = 2 x 10-7M), does not recognize PP but cross-reacts weakly with PYY. Antibodies NPY03 and NPY05 are IgG2 with respective Ka's of 6.7 x 109 and 2.5 x 1010 liters/mol. They interact with a C-terminal epitope on NPY (NPY 27-34, IC50 = 2 x 10-9 M for NPY03 and NPY 32-36, IC50 = 1 x 10-9 M for NPY05). These two antibodies cross-react with PYY whereas only NPY05 binds PP. NPY05 is unable to bind the free acid form of neuropeptide Y. The 32-36 COOH free subpeptide is recognized 50,000 less efficiently by NPY05 than its amidated form. Ant...