title: Rapid assembly of sensitive antigen-capture assays for Marburg virus, using in vitro selection of llama single-domain antibodies, at biosafety level 4.
abstract: There is a pressing need for rapid and reliable approaches to the delivery of sensitive yet rugged diagnostic assays specific for emerging viruses, to hasten containment of outbreaks when and wherever they occur. Within 3 weeks, we delivered an antigen-capture assay for Marburg virus (MARV) that was based on llama single-domain antibodies (sdAbs) selected at biosafety level 4. Four unique sdAbs were capable of independently detecting MARV variants Musoke, Ravn, and Angola without cross-reactivity with the 4 Ebola virus species. The unoptimized assays could be performed in <30 min and, at best, provided a visual read of 10-100 pfu in a 100-microL sample when a colorimetric substrate was used and 0.1-1 pfu when a chemiluminescent substrate was used. All the sdAbs were specific for nucleoprotein, with an assay sensitivity that was reliant on detergent-mediated exposure of polyvalent antigen. Our strategy highlights the potential of direct antibody selection on filoviruses as a guide for effective and fast diagnostic development.
authors.affiliation: Department of Virology and Immunology, Southwest Foundation for Biomedical Research, San Antonio, TX 78245, USA.
authors.forename: Laura J
authors.initials: LJ
authors.lastname: Sherwood
authors.forename: Lisa E
authors.initials: LE
authors.lastname: Osborn
authors.forename: Ricardo
authors.initials: R
authors.lastname: Carrion
authors.forename: Jean L
authors.initials: JL
authors.lastname: Patterson
authors.forename: Andrew
authors.initials: A
authors.lastname: Hayhurst
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood / Q000302:isolation & purification; D000956:Antigens, Viral / Q000302:isolation & purification*; D002161:Camelids, New World / Q000821:virology; D029043:Ebolavirus / Q000276:immunology / Q000302:isolation & purification; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D029024:Marburgvirus / Q000276:immunology* / Q000302:isolation & purification*; D012449:Safety; D012680:Sensitivity and Specificity
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral