Antibodies to peptide determinants in transforming growth factor .beta. and their applications

Abstract

Polyclonal antibodies have been raised to a series of synthetic peptides which correspond to essentially all regions of the transforming growth factor .beta.1 (TGF-.beta.1) molecule. All antisera were evaluated for their abilities to react with TGF-.beta.1 and TGF-.beta.2 in either the native or reduced form in enzyme-linked immunosorbent assays, Western blots, and immunoprecipitation assays. While all antisera demonstrated some ability to recognize TGF-.beta.1 in these systems, there was limited cross-reactivity with TGF-.beta.2, suggesting that substantial sequence of conformational differences exist between the two growth factors. On Western blots 5-10 ng of purified human platelet TGF-.beta.1 could be detected when probed with affinity-purified peptide antisera generated against peptides corresponding to residues 48-77, 50-75, and 78-109 of the 112 amino acid TGF-.beta.1 monomer. Antisera raised against peptides 50-75 and 78-109 were most effective in immunoprecipitating reduced and native 125I-TGF-.beta.1, respectively. The antisera also were tested for their effectiveness in blocking the binding of 125I-TGF-.beta.1 to its receptor. Anti-peptide 78-109 and anti-peptide 50-75 blocked 80% and 40% of the binding, respectively, while antibodies against amino-terminal peptides were without effect. These data suggest that the carboxyl-terminal region of TGF-.beta.1 may play a significant role in the binding of the native ligand to its receptor.