Myasthenia Gravis (MG) is a chronic autoimmune disorder affecting the postsynaptic neuromuscular junction (NMJ), where autoantibodies impair communication between nerves and skeletal muscles, leading to muscle weakness and fatigue. MG typically affects voluntary muscles, especially those controlling the eyes, face, and throat. Clinically, it can be categorized into three acquired forms: ocular MG (OMG), bulbar predominant MG (BMG), and generalized MG (GMG).
The disease is diagnosed through thorough clinical evaluation and detection of autoantibodies targeting specific components of the NMJ. These autoantibodies interfere with neuromuscular signaling by binding to or destroying receptors necessary for transmitting signals from the brain to the muscles. The presence and type of these antibodies not only assist in diagnosis but are also critical in guiding treatment plans.
In 80–90% of adults with generalized MG, autoantibodies target the acetylcholine receptor (AChR Ab). An additional 5–10% of patients have antibodies against muscle-specific tyrosine kinase (MuSK Ab). Among the remaining 10–15% of MG patients, whose sera are negative for both AChR and MuSK antibodies, the condition is classified as double-seronegative MG (dSN-MG). In these cases, autoantibodies may be present against other proteins such as low-density lipoprotein receptor-related protein 4 (LRP4), agrin, or cortactin, though detection rates vary based on ethnic and geographic factors.
To improve diagnostic sensitivity, several advanced testing methodologies have been introduced. Since 1984, BCNI has offered the gold-standard AChR RIPA assay. In 2017, we launched a more sensitive live cell-based assay (CBA) for AChR Abs, licensed from Oxford University and accredited by DAP/ISO, CAP, and CLIA. This live CBA has demonstrated higher sensitivity compared to fixed CBA, RIPA, and ELISA. Additionally, we developed a highly sensitive MuSK Ab assay using surface plasmon resonance (SPR), which complements the traditional MuSK RIPA assay. Through international collaborations, BCNI has also validated and implemented an LRP4 Ab CBA, enabling better detection in dSN-MG cases.
Testing Algorithm
BC Neuroimmunology employs an orthogonal algorithmic approach to testing for myasthenia gravis. In the event of an assay yielding negative results in a patient suspected of being positive for MG, additional testing with higher precision is performed on a confirmatory basis to ensure the highest degree of confidence in diagnosis.
Anti-AchR Ab Testing by Radioimmunoprecipitation
Our first pass myasthenia gravis test employs a radioimmunoprecipitation assay (RIPA) to assess presence of MG-related autoantibodies. We compare the sera of test patients to control subjects via a double antibody precipitation technique that employs the use of the Kronus antigen, a genetically engineered acetylcholine receptor. The high frequency of anti-AChR antibodies present in the sera of subtype-AChR Ab myasthenia gravis patients makes it an extremely reliable diagnostic.
BC Neuroimmunology offers two AchR Ab tests by RIPA: QUALITATIVE & QUANTITATIVE. All samples found to be POSITIVE for acetylcholine antibodies following a qualitative assay are repeated using the quantitative assay to permit comparison of titres.
Anti-MuSK Ab Testing by Radioimmunoprecipitation Assay
Muscle-specific tyrosine kinase (MuSK) is a protein involved in the formation and maintenance of the neuromuscular junction. It is activated by the ligand agrin and induces clustering of acetylcholine receptors on the post-synaptic muscle fibre.
Amongst AchR Ab-negative myasthenia gravis patients, MuSK autoantibodies are responsible for the largest representation of cases. Anti-MuSK MG is characterized by more prominent bulbar features (affecting nerves innervating muscles to do with the face, throat, and neck) and more frequent myasthenic crises.
Our anti-MuSK Ab test employs radioimmunoprecipitation assay (RIPA) to assess the presence of anti-MuSK antibodies. Only samples negative for the AChR Ab test will go on to be tested for anti-MuSK antibodies. Samples from patients that do not test positive for either AChR or MuSK antibodies are deemed double-seronegative.
As per MSC guidelines, MuSK Ab testing may only be requested by neurologists, ophthalmologists, and neuro-ophthalmologists in order to bill testing to provincial health services.
Anti-AChR Ab Testing by Live Cell-Based Assay
Live cell-based assays (CBA) are considered the most effective detection assay for antibody detection in myasthenia gravis and show excellent specificity and superior sensitivity when compared to other assay techniques. A significant proportion of MG sufferers (~10%) do not demonstrate presence of AChR or MuSK auto-antibodies (i.e. double seronegative patients) by the standard assay techniques. Cell-based assays have demonstrated the ability to detect AChR antibodies in 16-60% of MG patients previous assessed to be seronegative, improving detection sensitivity particularly in patients with milder disease presentation and in children.
The AchR Ab CBA can also be used to confirm results for samples that are false positive by AchR Ab RIPA. If a sample shows high non-specific binding via RIPA, the lab then tests the sample by CBA in order to provide a definitive result.
Anti-LRP4 Ab
Low density lipoprotein receptor-related protein 4 (LRP4) is a receptor protein on the neuromuscular junction that binds with the protein agrin and is critical for NMJ formation and maintenance via MuSK activation. Autoantibodies for LRP4 have been shown to be present and causal for MG in patients found to be double seronegative for AChR Ab and MuSK Ab.