Hepojoki Satu, Nurmi Visa, Vaheri Antti, Hedman Klaus, Vapalahti Olli, Hepojoki Jussi
Department of Virology, Haartman Institute, University of Helsinki, Helsinki, Finland.
Department of Virology, Haartman Institute, University of Helsinki, Helsinki, Finland; Helsinki University Central Hospital, Hospital District of Helsinki and Uusimaa, Laboratory Services, HUSLAB, Helsinki, Finland.
PLoS One. 2014 Sep 2;9(9):e106432. doi: 10.1371/journal.pone.0106432. eCollection 2014.
Chelated lanthanides such as europium (Eu) have uniquely long fluorescence emission half-lives permitting their use in time-resolved fluorescence (TRF) assays. In Förster resonance energy transfer (FRET) a donor fluorophore transfers its emission energy to an acceptor fluorophore if in sufficiently close proximity. The use of time-resolved (TR) FRET minimizes the autofluorescence of molecules present in biological samples. In this report, we describe a homogenous immunoassay prototype utilizing TR-FRET for detection of antibodies in solution. The assay is based on labeled protein L, a bacterial protein that binds to immunoglobulin (Ig) light chain, and labeled antigen, which upon association with the same Ig molecule produce a TR-FRET active complex. We show that the approach is functional and can be utilized for both mono- and polyvalent antigens. We also compare the assay performance to that of another homogenous TR-FRET immunoassay reported earlier. This novel assay may have wide utility in infectious disease point-of-care diagnostics.
诸如铕（Eu）等螯合镧系元素具有独特的长荧光发射半衰期，这使得它们能够用于时间分辨荧光（TRF）分析。在荧光共振能量转移（FRET）中，如果供体荧光团与受体荧光团足够接近，供体荧光团会将其发射能量转移给受体荧光团。使用时间分辨（TR）FRET可将生物样品中存在的分子的自发荧光降至最低。在本报告中，我们描述了一种利用TR-FRET检测溶液中抗体的均相免疫分析原型。该分析基于标记的蛋白L（一种与免疫球蛋白（Ig）轻链结合的细菌蛋白）和标记的抗原，它们与同一Ig分子结合后会产生具有TR-FRET活性的复合物。我们表明该方法是有效的，可用于单价和多价抗原。我们还将该分析性能与之前报道的另一种均相TR-FRET免疫分析的性能进行了比较。这种新型分析方法在传染病即时诊断中可能具有广泛的应用。
