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Co(III)-NTA Mediated Antigen Immobilization on a Fiber Optic-SPR Biosensor for Detection of Autoantibodies in Autoimmune Diseases: Application in Immune-Mediated Thrombotic Thrombocytopenic Purpura

Abstract : Autoantibodies are key biomarkers in clinical diagnosis of autoimmune diseases routinely detected by enzyme-linked immunosorbent assays (ELISAs). However, the complexity of these assays is limiting their use in routine diagnostics. Fiber optic-surface plasmon resonance (FO-SPR) can overcome these limitations, but improved surface chemistries are still needed to guarantee detection of autoantibodies in complex matrices. In this paper, we describe the development of an FO-SPR immunoassay for the detection of autoantibodies in plasma samples from immune-mediated thrombotic thrombocytopenic purpura (iTTP) patients. Hereto, hexahistidine-tagged recombinant ADAMTS13 (rADAMTS13-His(6)) was immobilized on nitrilotriacetic acid (NTA)-coated FO probes chelated by cobalt (Co(III)) and exposed to anti-ADAMTS13 autoantibodies. Initial studies were performed to optimize rADAMTS13-His 6 immobilization and to confirm the specificity of the immunoassay for detection of anti-ADAMTS13 autoantibodies with FO-SPR. The performance of the immunoassay was then evaluated by comparing Co(III)- and nickel (Ni(II))NTA stabilized surfaces, confirming the stable immobilization of the antigen in Co(III)-NTA-functionalized FO probes. A calibration curve was prepared with a dilution series of a cloned human anti-ADAMTS13 autoantibody in ADAMTS13-depleted plasma resulting in an average interassay coefficient of variation of 7.1% and a limit of detection of 0.24 ng/mL. Finally, the FO-SPR immunoassay was validated using seven iTTP patient plasma samples, resulting in an excellent correlation with an in-housedeveloped ELISA (r = 0.973). In summary, the specificity and high sensitivity in combination with a short time-to-result (2.5 h compared to 4-5 h for a regular ELISA) make the FO-SPR immunoassay a powerful assay for routine diagnosis of iTTP and with extension for any other autoimmune disease.
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https://hal.inrae.fr/hal-03155879
Contributor : Nelly Lucas <>
Submitted on : Tuesday, March 2, 2021 - 10:20:56 AM
Last modification on : Wednesday, March 3, 2021 - 3:14:13 AM

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Sara Horta, Jia-Huan Qu, Charlotte Dekimpe, Quintijn Bonnez, Aline Vandenbulcke, et al.. Co(III)-NTA Mediated Antigen Immobilization on a Fiber Optic-SPR Biosensor for Detection of Autoantibodies in Autoimmune Diseases: Application in Immune-Mediated Thrombotic Thrombocytopenic Purpura. Analytical Chemistry, American Chemical Society, 2020, 92 (20), pp.13880-13887. ⟨10.1021/acs.analchem.0c02586⟩. ⟨hal-03155879⟩

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