As reported by Rodero et al.26 (link), a Simoa interferon alpha (IFN-α) assay was developed using a Quanterix Homebrew Simoa assay and two autoantibodies specific for IFN-α isolated and cloned from two APS1/APECED patients recently described24 (link). The 8H1 antibody clone was used as a capture antibody after coating on 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 limits of detection (LOD) were calculated by the mean value of all blank runs + 3SDs and was 0.23 fg/mL.
Rodero M.P., Tesser A., Bartok E., Rice G.I., Della Mina E., Depp M., Beitz B., Bondet V., Cagnard N., Duffy D., Dussiot M., Frémond M.L., Gattorno M., Guillem F., Kitabayashi N., Porcheray F., Rieux-Laucat F., Seabra L., Uggenti C., Volpi S., Zeef L.A., Alyanakian M.A., Beltrand J., Bianco A.M., Boddaert N., Brouzes C., Candon S., Caorsi R., Charbit M., Fabre M., Faletra F., Girard M., Harroche A., Hartmann E., Lasne D., Marcuzzi A., Neven B., Nitschke P., Pascreau T., Pastore S., Picard C., Picco P., Piscianz E., Polak M., Quartier P., Rabant M., Stocco G., Taddio A., Uettwiller F., Valencic E., Vozzi D., Hartmann G., Barchet W., Hermine O., Bader-Meunier B., Tommasini A, & Crow Y.J. (2017). Type I interferon-mediated autoinflammation due to DNase II deficiency. Nature Communications, 8, 2176.
Other organizations :
Inserm, University of Trieste, University of Bonn, Manchester Academic Health Science Centre, University of Manchester, BIOASTER, Institut Pasteur, Université Paris Cité, Centre National de la Recherche Scientifique, Délégation Paris 5, Sorbonne Paris Cité, Institut des Maladies Génétiques Imagine, Istituto Giannina Gaslini, Hôpital Necker-Enfants Malades, Assistance Publique – Hôpitaux de Paris, IRCCS Materno Infantile Burlo Garofolo, Université Paris-Saclay, Université Paris-Sud, German Center for Infection Research
Coating of paramagnetic beads with 8H1 antibody clone (0.3 mg/mL)
Biotinylation of 12H5 antibody (biotin/Ab ratio = 30/1)
dependent variables
Concentration of interferon alpha (IFN-α) detected by the Simoa assay
control variables
Use of recombinant IFN-α17/αI (PBL Assay Science) as a standard curve after cross-reactivity testing
controls
Positive control: Recombinant IFN-α17/αI (PBL Assay Science) used as a standard curve
Negative control: Blank runs used to calculate the limit of detection (LOD)
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