Multicentre comparison of a diagnostic assay: aquaporin-4 antibodies in neuromyelitis optica

Abstract

ObjectiveAntibodies to cell surface central nervous system proteins help to diagnose conditions which often respond to immunotherapies. The assessment of antibody assays needs to reflect their clinical utility. We report the results of a multicentre study of aquaporin (AQP) 4 antibody (AQP4-Ab) assays in neuromyelitis optica spectrum disorders (NMOSD).MethodsCoded samples from patients with neuromyelitis optica (NMO) or NMOSD (101) and controls (92) were tested at 15 European diagnostic centres using 21 assays including live (n=3) or fixed cell-based assays (n=10), flow cytometry (n=4), immunohistochemistry (n=3) and ELISA (n=1).ResultsResults of tests on 92 controls identified 12assays as highly specific (0–1 false-positive results). 32 samples from 50 (64%) NMO sera and 34 from 51 (67%) NMOSD sera were positive on at least two of the 12 highly specific assays, leaving 35 patients with seronegative NMO/spectrum disorder (SD). On the basis of a combination of clinical phenotype and the highly specific assays, 66 AQP4-Ab seropositive samples were used to establish the sensitivities (51.5–100%) of all 21 assays. The specificities (85.8–100%) were based on 92 control samples and 35 seronegative NMO/SD patient samples.ConclusionsThe cell-based assays were most sensitive and specific overall, but immunohistochemistry or flow cytometry could be equally accurate in specialist centres. Since patients with AQP4-Ab negative NMO/SD require different management, the use of both appropriate control samples and defined seronegative NMOSD samples is essential to evaluate these assays in a clinically meaningful way. The process described here can be applied to the evaluation of other antibody assays in the newly evolving field of autoimmune neurology.

Large-Scale in-House Cell-Based Assay for Evaluating the Serostatus in Patients with Neuromyelitis Optica Spectrum Disorder Based on New Diagnostic Criteria

“…However, these assays reportedly have varying sensitivities (33–90%) 10. In the present fully blinded study, our in-house CBA detected AQP4-IgG in 80% of definite-NMOSD patients and 76% of high risk NMOSD patients, with 100% specificity.…”

“…An international multicenter study found that the commercial CBA kit (EUROIMMUN, Luebeck, Germany) had a specificity of 100% and sensitivities of between 51.5% and 86.4% 10. A similar study found that the CBA using live cells expressing human M23-AQP4 was more sensitive than the CBA using fixed cells 8.…”

Neuromyelitis optica presenting as acute bilateral ptosis

“…However, anti-aquaporin-4-IgG is highly specific for NMOSD: the Oxford laboratory's cell-based assay has a specificity of 99% 8 9. The absence of cerebrospinal fluid oligoclonal bands is consistent with this diagnosis and mild-to-moderate pleocytosis (range 6–380/µL (≤5)) may also occur in NMOSD 10…”

Expanding the range of immunopathology in neuromyelitis optica spectrum disorder

“…While NMO-IgG detection assays employ a variety of methodologies that vary in sensitivity and specificity, the Euroimmun cell-based assay (CBA) is observed to have 100% specificity for the diagnosis of NMO, in contrast to assays that use flow cytometry and live cells which return occasional false positive results 2. While this case demonstrates some laboratory and pathological features of multiple sclerosis (CSF-specific oligoclonal bands and active demyelination), these observations could potentially challenge the notion that NMO-IgG testing with the CBA is 100% specific and it should be noted that it is unlikely that any biological assay has 100% specificity.…”

“…In majority of cases an antibody that is reactive with aquaporin 4 (AQP4) (NMO-IgG) can be detected in the serum of the affected individuals. NMO-IgG has an extremely high sensitivity and specificity to the extent that NMO-IgG can be considered to be pathognomonic for the diagnosis of NMO 1 2. Moreover, immunopathological and in vitro studies are suggestive of a pathogenic role for NMO-IgG 3 4…”