Antiparanodal antibodies and IgG subclasses in acute autoimmune neuropathy

(A) Results of anti–contactin-1 and anti–Caspr-1 ELISA with baseline sera and subclass-specific IgG1-4 autoantibodies (grayscale) are shown as % of the OD of the total IgG (using anti-human IgG autoantibody) on y-axis. In patient 1, ELISA IgG subclasses were not measurable. ELISA revealed mainly IgG2 subclass in patient 2 (GBS), IgG2/IgG4 in patient 3 (A-CIDP), and IgG2/3 in patients 4 and 5 (A-CIDP). (B) Acute phase teased fibers subclass analysis with baseline sera showed binding to the paranodal regions (arrows) only when using IgG3-specific secondary antibody in patients 1 (GBS) and 4 (A-CIDP), visualized by photomicrographs of single immunofluorescence staining on teased fibers. (C) Anti–Caspr-1 IgG subclass ELISA with follow-up serum of patient 4 now revealed IgG 2/4 subclass. Photomicrographs show immunofluorescence staining of murine teased fibers with subclass-specific IgG3 and IgG4 autoantibodies at follow-up. Paranodal binding disappeared using IgG3 antibody, but occurred using IgG4 subclass-specific antibody, proving subclass switch in patient 4. Scale bar = 10 µm. A-CIDP = acute-onset chronic inflammatory demyelinating polyradiculoneuropathy; GBS = Guillain-Barré syndrome.

(A) Contactin-1 IgG ELISA of all patients at first assessment, of 66/161 patients at follow-up, and of 40 healthy controls. Optical density (OD) at 450 nm is displayed on the y-axis. The threshold for positive results is set at 5 SDs above the mean of controls (0.615). Patients 2–4 show positive results at first assessment. The other patients and controls show values below the threshold. Sera of patients 2, 4, and 5 are negative in anti–contactin-1 IgG ELISA at follow-up, as well as 63 other follow-up sera. Patients 1 and 3 were lost to follow-up. (B) Overlay photomicrographs of teased fibers double immunofluorescence and (C) contactin-1– and Caspr-1 (CNTNAP1)–transfected HEK293 cells show loss of anti–contactin-1/Caspr-1 autoantibodies in patient 2 (GBS) at follow-up (B.a, C.a, C.d), but colocalization of commercial anti–Caspr-1 antibody and sera at paranodes in patients 4 and 5 with A-CIDP (B.b, B.c, arrows). Both sera bind to Caspr-1 (CNTNAP1)–transfected HEK293 cells (C.e, C.f, arrowheads), but not to contactin-1–transfected cells (C.b, C.c). Scale bar = 10 µm. A-CIDP = acute-onset chronic inflammatory demyelinating polyradiculoneuropathy; GBS = Guillain-Barré syndrome; HEK = human embryonic kidney.