title: Production and characterization of monoclonal antibodies sensitive to conformation in the 5HT2c serotonin receptor.
abstract: mAbs that are sensitive to protein conformation can be helpful in studies of protein structure and function; in particular, mAb fragments are useful reagents in membrane protein crystallization. We immunized mice with the rat 5HT2c serotonin receptor and derived clonal hybridoma cells, which we tested for specific antigen reactivity by using the complementarity of purified protein from bacteria and receptor-embedded mammalian cell membranes. Nine mAbs met our criteria for specificity, affinity, and sensitivity to conformational features. Epitopes were mapped in various additional tests. Five of the nine mAbs have cytoplasmic epitopes, and two of these are sensitive to the ligand state of the receptor. These properties should be useful both for structural analysis and in probes of function.
authors.affiliation: Department of Biochemistry and Molecular Biophysics and the Howard Hughes Medical Institute, Columbia University, New York, NY 10032, USA.
authors.forename: Filippo
authors.initials: F
authors.lastname: Mancia
authors.forename: Susan
authors.initials: S
authors.lastname: Brenner-Morton
authors.forename: Risa
authors.initials: R
authors.lastname: Siegel
authors.forename: Zahra
authors.initials: Z
authors.lastname: Assur
authors.forename: Yonghua
authors.initials: Y
authors.lastname: Sun
authors.forename: Ira
authors.initials: I
authors.lastname: Schieren
authors.forename: Monica
authors.initials: M
authors.lastname: Mendelsohn
authors.forename: Richard
authors.initials: R
authors.lastname: Axel
authors.forename: Wayne A
authors.initials: WA
authors.lastname: Hendrickson
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D002462:Cell Membrane / Q000378:metabolism; D002469:Cell Separation; D003460:Crystallization; D003593:Cytoplasm / Q000378:metabolism; D018604:Epitope Mapping; D004926:Escherichia coli / Q000378:metabolism; D006825:Hybridomas / Q000378:metabolism; D008024:Ligands; D051379:Mice; D041681:NIH 3T3 Cells; D011487:Protein Conformation; D051381:Rats; D011985:Receptors, Serotonin / Q000737:chemistry*
chemical_list: D000911:Antibodies, Monoclonal; D008024:Ligands; D011985:Receptors, Serotonin