Quantitative Profiling of Nanoscopic Protein Aggregates Reveals Specific Fingerprint of TDP‐43‐Positive Assemblies in Motor Neuron Disease

Abstract Abnormal aggregation of TAR DNA‐binding protein 43 (TDP‐43) is a pathological hallmark of motor neuron disease (MND), yet current methods for quantifying these aggregates in biological samples remain limited in sensitivity and resolution. Here, single‐molecule fluorescence microscopy is applied to post‐mortem brain extracts to quantitatively characterize aggregates containing TDP‐43 at the individual particle level. The resulting aggregate fingerprints, consisting of morphological and compositional profiles, are sufficient to distinguish MND donors from neurologically normal controls and further discriminate between clinically distinct MND subgroups. Comparative proteomic analysis confirms and extends these findings, revealing convergent and complementary molecular signatures. These results demonstrate, for the first time, that single‐molecule aggregate profiling can stratify MND cases using patient‐derived tissues, paving the way for the development of sensitive minimally invasive diagnostics and mechanistically informed disease monitoring tools.