In clinical diagnostics, the accurate and sensitive detection of specific biomolecules is vital for effective disease diagnosis and monitoring. This is commonly achieved through the use of immunoassays, an approach which relies on highly specific interactions between antibodies and antigens, often coupled to spectroscopic tools to facilitate biomarker detection and quantification. However, these spectroscopic approaches experience inherent limitations, particularly in the context of multiplexing experiments, where the overlap of broad emission ranges can lead to interference when attempting to analyse several biomarkers simultaneously. This issue may be resolved by instead coupling immunoassay approaches to mass spectrometry (MS), where the detection of a particular m/z value, produced by the cleavage of a mass tag from biomolecule of interest may be used to indicate the presence of antigen. In this approach, the discrete nature of m/z values allows for the detection of a range of biomolecules in a single experiment with limited interference.
In this work, two novel immunoassay designs are explored, focusing on developing methods that improve sensitivity and simplicity of detection, in an attempt to advance MS-immunoassays as potential diagnostic tools. In the first approach, an amplified MS-immunoassay suitable for matrix assisted laser desorption ionisation (MALDI-MS) detection of low-abundance biomarkers is proposed. As part of this work, two photocleavable mass tags were synthesised and shown to exhibit the desired photocleavable properties under conditions present in MALDI-MS. In an alternative approach, a paper substrate-based MS-immunoassay is presented, relying on a mass tag able to be cleaved under the conditions present in electrospray ionisation MS. In each design, preliminary studies have demonstrated detection of selected analytes, however further optimisation still required. Further studies hope to improve the accuracy, sensitivity and multiplexing capabilities of these approaches and employ these mass spectrometric immunoassays as powerful diagnostic tools in a range of diseases.