Differential inhibition of receptor activation by two mouse monoclonal antibodies specific for the human leukotriene B4 receptor, BLT1

• Published in: International immunopharmacology Vol. 3; no. 13-14; pp. 1829 - 1839
• Main Authors: SABIRSH, A, PETTERSSON, A, BOKETOFT, A, KOTARSKY, K, OWMAN, C
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier 01.12.2003
• Subjects: Animals; Antibodies, Monoclonal - biosynthesis; Antibodies, Monoclonal - immunology; Antibody Specificity; Antigen-Antibody Reactions; Binding, Competitive; Biological and medical sciences; Calcium - metabolism; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Fluorescence Polarization; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Gene Expression; General aspects; HeLa Cells; Humans; Leukotriene Antagonists; Leukotriene B4 - biosynthesis; Leukotriene B4 - immunology; Luciferases - metabolism; Mice; Molecular immunology; Receptors, G-Protein-Coupled - metabolism; Receptors, Leukotriene B4 - biosynthesis; Receptors, Leukotriene B4 - immunology; Receptors, Leukotriene B4 - physiology; Human; Binding; Antigenic determinant; Enzyme; Rodentia; Monoclonal antibody; Chemotaxis; Cell surface; cell surface; Epitope mapping; Binding; Enzyme-linked immunosorbent assay; Chemotaxis; Vertebrata; Mammalia; Immunosorbent; Leukotriene B4; Mouse; Receptor; Animal; Differential method; Biological receptor
• ISSN: 1567-5769; 1878-1705
• DOI: 10.1016/j.intimp.2003.08.009

The inflammatory mediator leukotriene B(4) (LTB(4)) binds to and activates a G-protein-coupled receptor named BLT(1). We have previously produced two monoclonal antibodies, named 7B1 and 14F11, that bind specifically to this receptor. Using a HeLa cell line expressing human BLT(1), we find that both antibodies inhibit LTB(4)-induced calcium release, and activation of a MAP-kinase-sensitive luciferase reporter system. The normal chemotactic movement of polymorphonuclear cells towards higher LTB(4) concentrations was also strongly inhibited by both antibodies. Neither antibody was found to activate BLT(1), and experiments using cyclic peptide fragments of the BLT(1) n-terminal and extracellular loops showed that these antibodies bind only to complex epitopes in the tertiary, membrane bound, conformation of the receptor protein. In ligand binding experiments, 7B1 was found to be a competitive antagonist, while 14F11 was a noncompetitive antagonist that inhibited receptor activation, but not agonist (LTB(4)) binding. 14F11 will be a useful tool for studying the mechanisms of receptor activation.