Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disorder that currently lacks validated molecular biomarkers for early diagnosis and prognosis, severely delaying personalized care. Interleukin 18 (IL-18), a proinflammatory cytokine linked to NLRP-3 inflammasome activation, has emerged as a promising biomarker for ALS. However, traditional colorimetric Enzyme-Linked Immunosorbent Assays (ELISAs) lack the sensitivity to distinguish IL-18 levels between Fast- and Slow-progressing ALS patients. To overcome this, we developed a highly sensitive electrochemical ELISA (e-ELISA) test by systematically optimizing key parameters, including the capture antibody immobilization strategy, the electrochemical mediator, and reagent concentrations. We then applied the optimized e-ELISA protocol to quantify IL-18 in 3D innervated skin models constructed using 3D-printed methacrylated hyaluronic acid (MeHA) and electrospun polylactic acid (PLLA) fibers, and colonized with patient-derived fibroblasts and neuronal cells. Reaching a limit of detection of 1.77 pg✕mL -1 , the e-ELISA not only differentiated ALS models from the healthy control but, most critically, distinguished between a Fast- and a Slow-progressing ALS models based on significantly different IL-18 concentrations. By discriminating IL-18 levels in biologically representative models, this work validates the developed high-performance e-ELISA for personalized clinical use, providing a foundation for the design of portable diagnostic devices. • A high-performance e-ELISA test was developed for precise ALS diagnosis. • BSA-crosslinked coating, with oriented covalent immobilization of antibodies, lowered the LOD. • The use of HQ as an electrochemical mediator improved assay sensitivity. • e-ELISA was validated on patient-derived 3D innervated skin models. • IL-18 levels can differentiate between Fast- and Slow-progressing ALS patients.
Furlan et al. (Sun,) studied this question.