Antipeptide antibodies of predetermined specificity recognize and neutralize the bioactivity of the pan-specific hemopoietin interleukin 3.
Ten peptides that corresponded to portions of the T cell lymphokine pan-specific hemopoietin interleukin 3 (IL 3) were synthesized, coupled to keyhole limpet hemocyanin, and used to raise antipeptide antibodies in rabbits. These antisera reacted to varying degrees with native biologically active IL 3. Antibodies directed against peptides corresponding to residues 1-29 at the NH2 terminus, 123-140 at the COOH terminus, and to residues 64-82 and 91-112 were affinity-purified on peptide columns. Immunoabsorbent columns produced from affinity-purified antibodies to the 1-29, 91-112, and 123-140 although not the 64-82 peptide were effective in depleting biologically active IL 3 from conditioned medium. However, the antibodies specific for peptides 91-112 and 123-140 had only a low affinity for native IL 3 and it was only in the case of the anti-1-29 antibodies that a significant amount of IL 3 remained bound after extensive washing and could be recovered from the column by acid elution. The affinity-purified antibodies directed to peptides 1-29, 91-112, and 123-140 significantly inhibited the biological activity of IL 3, although with different dose-response characteristics. Anti-1-29 antibodies inhibited bioactivity over a wide range of concentrations (down to 20 ng/ml) although the inhibition was never complete. In contrast, the anti-91-112 antibodies, although effective only at high concentrations, produced complete inhibition of biological activity. These experiments demonstrated that antibodies to defined peptides can be used to generate antibodies to native IL 3 and should form useful tools in analyzing the structure and function of the native molecules.
D000818:Animals; D000906:Antibodies / Q000276:immunology*; D000918:Antibody Specificity; D002846:Chromatography, Affinity; D000939:Epitopes / Q000276:immunology; D007377:Interleukin-3 / Q000276:immunology* / Q000494:pharmacology; D009500:Neutralization Tests; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology*; D011817:Rabbits
D000906:Antibodies; D000939:Epitopes; D007377:Interleukin-3; D010446:Peptide Fragments