Detection and characterisation of the human myelin oligodendrocyte glycoprotein antibody response

Autoantibodies targeting myelin oligodendrocyte glycoprotein (MOG Ab) have become an important biomarker for a novel entity of central nervous system demyelinating syndromes. As the clinical relevance of MOG Ab is becoming increasingly clear, an in-depth characterisation of the human MOG Ab response is warranted. This thesis aims to improve MOG Ab detection and characterises key antibody features of epitope, binding sensitivity, and isotype, in the largest cohort of MOG Ab-positive patients. Flow cytometry cell-based assays (CBA) preserves the native MOG structure and was used to detect and quantify MOG Ab binding. A blinded study using a fixed commercial assay and paraformaldehyde-fixed CBA showed a significant loss of MOG Ab seropositivity and reduced sensitivity, shedding light on the importance of antigen conformation for accurate MOG Ab detection. The MOG Ab response is predominately IgG1, produced peripherally, and elevated titres were observed in more severe phenotypes and during active disease. Proline42 was the immunodominant epitope in human MOG Ab responses, followed by Histidine103/Serine104. Reactivity toward these extracellular epitopes were stable over time, and compared to children, adult responses were more polyclonal, and epitope specificity broadened with age. Interestingly, adults who did not bind Proline42 were likely to relapse, a feature which could be harnessed to predict a relapsing course. Published analyses were applied to a flow cytometry dataset to investigate influences MOG Ab seropositivity. A binding ratio was subject to the non-specific background binding inherent to serum, which reduced assay sensitivity. Flow cytometry CBA is a robust and highly sensitive method to detect MOG Ab and we established guidelines to standardise flow cytometry MOG Ab analyses. The results presented in this thesis has led to major advances in MOG Ab detection, patient diagnosis, prognosis, and our understanding of the disease aetiology