title: Rapid homogeneous immunoassay based on time-resolved Förster resonance energy transfer for serodiagnosis of acute hantavirus infection.
abstract: We recently introduced a homogeneous immunoassay based on time-resolved Förster resonance energy transfer (TR-FRET) elicited by fluorophore-labeled antigen and fluorophore-labeled protein L, bound by an immunoglobulin. As the first clinical application, we employ this approach (LFRET) in serodiagnosis of Puumala hantavirus (PUUV) infection. A reference panel containing serum from individuals with acute (n = 21) or past (n = 17) PUUV infection and from PUUV-seronegative individuals (n = 20) was used to define the parameters. The clinical assay performance was evaluated with a prospectively collected serum panel (panel 2; n = 153). Based on the results for panel 1, the threshold for positivity was set at a signal level that was 3-fold over background, while those with a signal <3-fold over the background level were considered PUUV seronegative. With panel 1, 20/21 acute- and 7/10 past-infection samples induced positive signals, compared to 0/20 seronegatives. With panel 2, a positive signal was obtained in 39/40 acute- and 4/10 past-infection samples, as opposed to 7/103 seronegatives. However, after IgG depletion, 58/61 acute-infection samples were LFRET positive, while all past-infection and seronegative samples were negative, corresponding to 100% specificity and 95% sensitivity in detection of acute PUUV infection. We demonstrate that the novel immunoassay is a promising tool for rapid serodiagnosis of acute Puumala virus infection.
authors.affiliation: Department of Virology, University of Helsinki, Helsinki, Finland.
authors.forename: Satu
authors.initials: S
authors.lastname: Hepojoki
authors.affiliation: Department of Virology, University of Helsinki, Helsinki, Finland jussi.hepojoki@helsinki.fi.
authors.forename: Jussi
authors.initials: J
authors.lastname: Hepojoki
authors.affiliation: Department of Virology, University of Helsinki and Helsinki University Hospital, Helsinki, Finland.
authors.forename: Klaus
authors.initials: K
authors.lastname: Hedman
authors.affiliation: Department of Virology, University of Helsinki and Helsinki University Hospital, Helsinki, Finland Department of Veterinary Biosciences, Faculty of Veterinary Medicine, University of Helsinki, Helsinki, Finland.
authors.forename: Olli
authors.initials: O
authors.lastname: Vapalahti
authors.affiliation: Department of Virology, University of Helsinki and Helsinki University Hospital, Helsinki, Finland.
authors.forename: Antti
authors.initials: A
authors.lastname: Vaheri
mesh_terms: D031541:Fluorescence Resonance Energy Transfer; D018778:Hantavirus Infections / Q000175:diagnosis*; D006801:Humans; D011446:Prospective Studies; D029262:Puumala virus / Q000276:immunology*; D012680:Sensitivity and Specificity; D012698:Serologic Tests / Q000379:methods*; D013997:Time Factors