Electronic Poster | Session 2

029 – Validating a flow cytometry live cell-based assay to detect myelin oligodendrocyte glycoprotein antibodies for clinical diagnostics

Joseph A. Lopez (1, 2) – Fiona Tea (1, 2) – Ganesha S.P. Liyanage (1, 2) – Alicia Zou (1, 2) – Deepti Pilli (1, 2) – Chelsea B. Bassett (1, 2) – Vera Merheb (1) – Fiona X. Z. Lee (1) – Selina Thomsen (1) – Sudarshini Ramanathan (1, 2) – David A. Brown (3) – Russell C. Dale (1, 2, 4) – Fabienne Brilot (1, 2, 4)
Brain Autoimmunity Group, Kids Neuroscience Centre, Kids Research at the Children’s Hospital at Westmead, Sydney, Australia (1) – Discipline of Child and Adolescent Health, The University of Sydney, Sydney, Australia (2) – New South Wales Health Pathology-ICPMR and Westmead Institute for Medical Research, The University of Sydney, Sydney, Australia (3) – Brain and Mind Centre, University of Sydney, Sydney, Australia (4)

Myelin oligodendrocyte glycoprotein (MOG) has been a candidate autoantigen in demyelinating disorders over the last 40 years. Currently, there is an increasing demand for the diagnostic testing of MOG antibodies which aids in discerning certain demyelinating disorders from multiple sclerosis. Flow cytometry live cell-based assays (CBAs) for high-throughput and quantitative MOG antibody detection have been reported in several studies. However, translation of these research assays to real-world diagnostics is lacking. Therefore, the aim of this study was to validate a MOG antibody flow cytometry live CBA for implementation in clinical diagnostics. A total of 1,248 adult patient sera were screened using a research-based flow cytometry live CBA involving ZsGreen+ MOG-expressing cells and ZsGreen+ control cells (double-green assay; n=613), and an optimised streamline assay involving ZsGreen+ MOG-expressing cells and mCherry+ control cells (red-green assay; n=635). The red-green assay effectively halves the experimental workload by combining antigen-positive and -negative cells to be tested together, making it more applicable in a diagnostic laboratory. In the red-green assay, 602/635 (94.8%) patient sera had a confirmed serostatus following two replicate experiments while 577/613 (94.1%) had a confirmed serostatus in the double-green assay. The inter-assay mean coefficient of variability of MOG antibody-positive sera was 4% lower in the red-green assay (57/635 MOG antibody-positive) compared to the double-green assay (61/613 MOG antibody-positive). These data suggest that a streamlined flow cytometry live CBA can be optimised for accredited diagnostic use with reliable serostatus determination and improved precision. Future implementation of this assay would fulfil increasing requests for MOG antibody testing to inform clinical diagnosis and treatment of MOG antibody-associated demyelination.