Neurodegenerative diseases (NDs) pose significant challenges to both longevity and quality of life, affecting millions worldwide and ranking as a major cause of global morbidity and disability. There is an urgent need for less invasive, cost-effective diagnostic methods that can reliably detect ND-related biomarkers in readily available biofluids such as serum. In response to this need, we have developed an ultra-sensitive assay that utilizes magnetic nanoparticles and a novel, tailor-made turn-on fluorescent probe named F-SPG. This innovative assay enables the detection of neurofilament light chain (NfL) at femtomolar concentrations, with a remarkable detection limit of 24 fM which is at least 18 times more sensitive than the conventional ELISA kits. Our assay's design eliminates the need for traditional secondary antibody in immunoassay, thereby streamlining the diagnostic process. Recoveries exceeding 95% underscore the assay's precision and reliability. It has proven effective in distinguishing Alzheimer's Disease patients from healthy individuals by quantifying serum NfL levels, showcasing its potential as a cost-effective, ultra-sensitive tool for the early screening of neurodegenerative diseases.