A Fluorescence Immunoassay for Serum Neurofilament Light Chain Detection Based on Magnetic Separation and Fluorescent Nanospheres Labeling

Neurofilament light chain protein (NfL) has emerged as a valuable biomarker in various neurodegenerative diseases (NDs). Initially, NfL detection relied on invasive cerebrospinal fluid (CSF) sampling. Although the concentration of NfL in serum is low, it shows a positive correlation with that in CSF. However, current methods for serum NfL detection suffer from limitations such as inadequate sensitivity and operational complexity. This study developed a novel assay leveraging antibody-conjugated magnetic nanospheres for target capture and fluorescent nanospheres for quantification, significantly enhancing the sensitivity, accuracy, and stability for serum NfL detection. Under optimal conditions, the method exhibited a linear relationship between fluorescence intensity and NfL concentration across 0.005–10 ng/mL with a detection limit of 1.4 pg/mL. Spiked recovery experiments performed in 5-fold diluted human serum samples demonstrated 95.4–103.2% recovery, confirming high accuracy and clinical utility. Overall, the established method may offer a promising new approach for the auxiliary diagnosis of NDs.