As previously reported (5 (link)), Simoa digital ELISA specific for IFN-α was developed using a Quanterix Homebrew Assay, and two autoantibodies specific for IFN-α were isolated and cloned from two APS1/APECED patients (59 (link)). The 8H1 antibody clone was used as a capture antibody after coating paramagnetic beads (0.3 mg/ml), and the 12H5 was biotinylated (biotin/Ab ratio = 30:1) and used as the detector. Recombinant IFN-α17/αI (PBL Assay Science) was used as a standard curve after cross-reactivity testing. The limit of detection was calculated by the mean value of all blank runs ± 3 SDs and was 0.23 fg/ml.
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Ultrasensitive IFN-α Digital ELISA
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Corresponding Organization : Sorbonne Paris Cité
Other organizations : Institut Pasteur, Inserm, University of Duisburg-Essen, Institut des Maladies Génétiques Imagine, Assistance Publique – Hôpitaux de Paris, Hôpital Necker-Enfants Malades, Hôpital Robert-Debré, Université de Strasbourg, Centre National de la Recherche Scientifique, Institut de Biologie Moléculaire et Cellulaire, Immungenetics (Germany), Institut Cochin, Institut de Recherche en Infectiologie de Montpellier, Université de Montpellier
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Variable analysis
independent variables
- Coating paramagnetic beads with 8H1 antibody clone (0.3 mg/ml)
- Biotinylating 12H5 antibody (biotin/Ab ratio = 30:1)
- Concentration of IFN-α detected using the Simoa digital ELISA
- Recombinant IFN-α17/αI (PBL Assay Science) used as a standard curve after cross-reactivity testing
- Recombinant IFN-α17/αI (PBL Assay Science) used as a standard curve
- Blank runs used to calculate the limit of detection (0.23 fg/ml)
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