title: Determining the specificity of monoclonal antibody HPT-101 to tau-peptides with optical tweezers.
abstract: Optical tweezers-assisted dynamic force spectroscopy is employed to investigate specific receptor-ligand interactions on the level of single binding events. In particular, we analyze binding of the phosphorylation-specific monoclonal antibody (mAb) HPT-101 to synthetic tau-peptides with two potential phosphorylation sites (Thr231 and Ser235), being the most probable markers for Alzheimer's disease. Whereas the typical interpretation of enzyme-linked immunosorbent assay (ELISA) suggests that this monoclonal antibody binds exclusively to the double-phosphorylated tau-peptide, we show here by DFS that the specificity of only mAb HPT-101 is apparent. In fact, binding occurs also to each sort of monophosphorylated peptide. Therefore, we characterize the unbinding process by analyzing the measured rupture force distributions, from which the lifetime of the bond without force τ0, its characteristic length xts, and the free energy of activation ΔG are extracted for the three mAb/peptide combinations. This information is used to build a simple theoretical model to predict features of the unbinding process for the double-phosphorylated peptide purely based on data on the monophosphorylated ones. Finally, we introduce a method to combine binding and unbinding measurements to estimate the relative affinity of the bonds. The values obtained for this quantity are in accordance with ELISA, showing how DFS can offer important insights about the dynamic binding process that are not accessible with this common and widespread assay.
authors.affiliation: Department of Experimental Physics I, Leipzig University , Linnéstraße 5, D-04103 Leipzig, Germany.
authors.forename: Tim
authors.initials: T
authors.lastname: Stangner
authors.forename: Carolin
authors.initials: C
authors.lastname: Wagner
authors.forename: David
authors.initials: D
authors.lastname: Singer
authors.forename: Stefano
authors.initials: S
authors.lastname: Angioletti-Uberti
authors.forename: Christof
authors.initials: C
authors.lastname: Gutsche
authors.forename: Joachim
authors.initials: J
authors.lastname: Dzubiella
authors.forename: Ralf
authors.initials: R
authors.lastname: Hoffmann
authors.forename: Friedrich
authors.initials: F
authors.lastname: Kremer
mesh_terms: D000544:Alzheimer Disease / Q000378:metabolism; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D000939:Epitopes / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D008962:Models, Theoretical; D052898:Optical Tweezers; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D013057:Spectrum Analysis; D013816:Thermodynamics; D016875:tau Proteins / Q000737:chemistry* / Q000276:immunology*
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D010455:Peptides; C000591032:monoclonal antibody HPT-101; D016875:tau Proteins