title: Label free checkerboard assay to determine overlapping epitopes of Ebola virus VP-40 antibodies using surface plasmon resonance.
abstract: Immunoassay formats, in which antibodies provide sensitivity and specificity, are often utilized to provide rapid and simple diagnostic tests. Surface plasmon resonance is frequently used to evaluate the suitability of antibodies by determining binding kinetics to agents or surrogate antigens. We used SPR to evaluate a number of commercial monoclonal antibodies as well as single domain antibodies produced in-house. All the antibodies targeted the Ebola virus viral protein 40 (VP40). We determined the ability of each antibody to bind to immobilized VP40, and ensured they did not bind Ebola glycoprotein or the nucleoprotein. A subset of the monoclonal antibodies was immobilized to characterize antigen capture in solution. It can be advantageous to utilize antibodies that recognize distinct epitopes when choosing reagents for detection and diagnostic assays. We determined the uniqueness of the epitope recognized by the anti-VP40 antibodies using a checkerboard format that exploits the 6×6 array of interactions monitored by the Bio-Rad ProteOn XPR36 SPR instrument. The results demonstrate the utility of surface plasmon resonance to characterize monoclonal and recombinant antibodies. Additionally, the analysis presented here enabled the identification of pairs of anti-VP40 antibodies which could potentially be utilized in sandwich type immunoassays for the detection of Ebola virus.
authors.affiliation: Naval Research Laboratory, Center for Biomolecular Science and Engineering, 4555 Overlook Ave SW, Washington, DC 20375, USA.
authors.forename: George P
authors.initials: GP
authors.lastname: Anderson
authors.affiliation: Naval Research Laboratory, Center for Biomolecular Science and Engineering, 4555 Overlook Ave SW, Washington, DC 20375, USA.
authors.forename: Jinny L
authors.initials: JL
authors.lastname: Liu
authors.affiliation: Naval Research Laboratory, Center for Biomolecular Science and Engineering, 4555 Overlook Ave SW, Washington, DC 20375, USA.
authors.forename: Dan
authors.initials: D
authors.lastname: Zabetakis
authors.affiliation: Naval Research Laboratory, Center for Biomolecular Science and Engineering, 4555 Overlook Ave SW, Washington, DC 20375, USA.
authors.forename: Patricia M
authors.initials: PM
authors.lastname: Legler
authors.affiliation: Naval Research Laboratory, Center for Biomolecular Science and Engineering, 4555 Overlook Ave SW, Washington, DC 20375, USA. Electronic address: ellen.goldman@nrl.navy.mil.
authors.forename: Ellen R
authors.initials: ER
authors.lastname: Goldman
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D000915:Antibody Affinity; D000918:Antibody Specificity; D000956:Antigens, Viral / Q000276:immunology* / Q000378:metabolism; D001666:Binding Sites, Antibody; D029043:Ebolavirus / Q000276:immunology* / Q000378:metabolism; D018604:Epitope Mapping / Q000379:methods*; D000939:Epitopes; D007118:Immunoassay / Q000379:methods*; D007700:Kinetics; D011485:Protein Binding; D011489:Protein Denaturation; D055550:Protein Stability; D061905:Single-Domain Antibodies / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D020349:Surface Plasmon Resonance; D013696:Temperature; D014763:Viral Matrix Proteins / Q000276:immunology* / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D000939:Epitopes; D061905:Single-Domain Antibodies; C080981:VP40 protein, virus; D014763:Viral Matrix Proteins
keywords: Binding kinetics; Ebola virus; Epitope; Monoclonal antibody; Single domain antibody; Surface plasmon resonance; VP40