Validation and optimisation of a commercial cell-based assay for detection of nodo-paranodal antibodies.

Abstract

A minority of patients fulfilling diagnostic criteria for chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) have been found to have autoantibodies against components of nodal and paranodal structures in peripheral nerves. These autoantibodies may confer distinct clinical features, including a lower likelihood of response to treatment with intravenous immunoglobulin (IVIg). There are currently limited options available for the detection of these autoantibodies in the diagnostic setting. We tested serum samples from 30 patients with CIDP, 40 disease controls [20 with myasthenia gravis and 20 with systemic lupus erythematosus (SLE)] and 52 healthy controls for the presence of antibodies against neurofascin 155 (NF155), neurofascin 186 (NF186), contactin-1 (CNTN1) and the contactin-1/contactin-associated protein 1 (CNTN1/CASPR1) complex using a commercial transfected HEK293 cell-based indirect immunofluorescence immunoassay (EUROImmun). We detected nodo-paranodal antibodies in six of 30 CIDP patients, including five who had previously tested positive for nodo-paranodal antibodies using enzyme-linked immunosorbent assay (ELISA). There was one positive test for anti-CNTN1 in a disease control with SLE and membranous glomerulonephritis, with no positive tests in healthy controls. We modified the manufacturer's staining protocol by using biotinylated anti-IgG and streptavidin-FITC labelling, which increased the analytical sensitivity of the assay. Our modified assay retained its robustness in the presence of interfering substances (haemolysed, lipaemic and icteric samples) and serum with high non-specific background immunofluorescent staining. We successfully modified and validated a commercial indirect immunofluorescence immunoassay for the qualitative detection of antibodies against NF155, NF186, CNTN1 and CNTN1/CASPR1. This could lead to more rapid diagnosis of patients with these autoantibodies, avoiding costly and ineffective treatments.