An immuno-DOT diagnostic assay for autoimmune nodopathy

Abstract

Objectives: Autoimmune nodopathy (AN) is a life-threatening peripheral neuropathy mediated by four autoantibodies targeting axoglial cell adhesion molecules at the nodes of Ranvier: Neurofascin-155 (Nfasc155), PanNeurofascin (PanNfasc), Contactin-1 (CNTN1), and Contactin-associated protein 1 (CASPR1). Antibody detection is a strong biomarker for AN diagnosis and treatment monitoring. The aim of this study was to develop an immuno-dot assay (immuno-DOT) compatible with routine implementation in medical laboratories. Methods: This new approach was compared to standard techniques: indirect immunofluorescence assay, cell-based assay, and ELISA. Sensitivities (Se) and specificities (Sp) were calculated on a cohort composed of 58 patients diagnosed with AN, 50 seronegative patients with chronic inflammatory demyelinating polyradiculoneuropathy, 20 healthy controls, 30 patients with Guillain-Barré syndrome, 20 with monoclonal gammopathy and 20 with Charcot-Marie-Tooth disease. The patients were diagnosed with AN based on compatible electro-clinical arguments and at least two positive standard techniques. Results: Immuno-DOT sensitivities and specificities were Se=91%, Sp=97% for anti-Nfasc155; Se=80%, Sp=94% for anti-PanNfasc; Se=93%, Sp=98% for anti-CNTN1; and Se=87%, Sp=94% for anti-CASPR1. Immuno-DOT allowed the diagnosis within 3h and the accurate follow-up of the immune reactivity and isotype, and dot intensity correlated with antibody titers following treatments. A longitudinal study indicated that immuno-DOT yielded reliable results even after six months of storage at-20°C. Conclusions: The diagnostic performance of immuno-DOT was satisfactory and compatible with routine implementation in medical laboratories.