Myasthenia Gravis (MG) is an autoimmune disorder that leads to weakness and diminished control over voluntary muscles. It results from reduced signaling in the neuromuscular junction (NMJ) that connects nerves to muscle fibers. Typically, the areas of reduced functioning are the muscles of the eyes, face, and throat. There exist three types of acquired MG: ocular (OMG), bulbar predominant (BMG), and generalized (GMG).
In MG, the body produces antibodies (Ab) that bind with receptors in the NMJ, blocking or destroying them, thereby preventing chemical signaling from the brain to the muscles.
The majority of cases (~85%) of MG are caused by autoantibodies targeting the acetylcholine receptors (AChR). Approximately 6% of cases are caused by autoantibodies targeting the extracellular domain of muscle specific tyrosine-kinase (MuSK) receptors. Amongst the ~15% of cases of MG that are found to be AChR Ab-negative, however, MuSK Abs have been reported in patients with frequencies ranging from 0 – 64%, depending on ethnic group and geographic location.
The remaining ~10% of cases are considered double-seronegative MG, as the patients’ sera appears negative for both AChR and MuSK antibodies. These cases can be caused by autoantibodies to a variety of other proteins, including but not limited to lipoprotein-related protein 4 (LRP4), agrin, and cortactin.
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 Surface Plasmon Resonance
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 surface plasmon resonance (SPR) technology to assess and quantify the presence of anti-MuSK antibodies. MuSK proteins are immobilized on a gold chip and the patient’s sera is passed over it. Binding is then measured based on the change in refractive index of light passing through the surrounding fluid. 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.
MuSK Ab testing may only be requested by neurologists, ophthalmologists, and neuro-ophthalmologists. Only samples found negative for anti-AChR Abs will go on to be tested for anti-MuSK Abs.
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.
Tests Available for Research Purposes Only
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. Accreditation for our anti-LRP4 CBA is pending.