Development and pre-qualification of a facile and ultra-sensitive LRRK2 LC/MS assay using high-affinity unmodified immobilization for immunocapture: meet bioanalytical challenges and search practical solutions

BACKGROUND: Quantifying low-abundance LRRK2 in CSF is analytically challenging but remains critical for Parkinson's disease biomarker development. METHODS: An immunocapture LC-MS assay using high-affinity unmodified antibody immobilization on Protein G beads coupled with a Vanquish Neo-Stellar nano-LC-MS platform was developed and pre-qualified. RESULTS: In 250 µL CSF, the assay achieved an LLOQ of 2 pg/mL, reduced antibody consumption (0.8 µg/sample), robust surrogate-to-matrix parallelism, and acceptable accuracy and precision under fit-for-purpose criteria. CONCLUSIONS: This practical, ultra-sensitive workflow overcomes key limitations of prior SISCAPA-based methods and is suitable for high-throughput translational and clinical LRRK2 biomarker studies. This flexible framework is extendable to other low-abundance protein biomarkers requiring rugged, automated, nano-LC mass spectrometric quantification in complex biological matrices.