To develop a highly sensitive and specific Surface Plasmon Resonance-based immunoassay for antibodies against Acetycholine Receptor that is paramount to Myasthenia Gravis clinical diagnosis

Myasthenia Gravis is the prototypical antibody-mediated autoimmune disease, with antibodies directed against the acetylcholine receptor at the postsynaptic motor endplate of the neuromuscular junction in 80–90% of patients. AChR MG is chiefly diagnosed by the detection of antibodies in patient sera using a radioimmunoprecipitation assay (RIPA) in which antibodies precipitate I-125-labelled AChR. Despite excellent sensitivities and specificities, RIPAs are limited by the use of radioactive labels. Thus novel label-free assays are attractive alternatives in clinical laboratory diagnoses. Surface Plasmon Resonance (SPR) detects biomolecular interactions in real time without the need for radio- or enzyme- labels. In SPR, the resultant sensorgrams that monitor binding interactions over time provide information on concentration, affinity, binding specificity and antibody isotypes. Most importantly, as it does not involve washing steps, SPR can detect both high and low affinity antibodies.

An SPR immunoassay was developed for AChR antibodies. Sera were pretreated with Protein A to obtain purified IgG which were flowed over a sensorchip composed of AChR captured by immobilized alpha-bungarotoxin. We analyzed the binding responses (antibody concentration) and further characterized the IgG subtypes. And in a separate assay using a low density surface of AChR, we determined the antibody dissociation rates, an indicator of antibody affinity.

We have thus far analysed 16 sera from MG patients previously tested as AChR antibody-positive and 10 healthy control sera. All 16 RIPA positive sera tested positive in the SPR immunoassay, whilst the 10 healthy control sera tested negative.

The SPR immunoassay is a comprehensive platform that provides information on the concentration, IgG isotype and affinity of AChR antibodies to better aid in the diagnosis of AcR antibody-mediated MG.