Microarray Chip-Based High-Throughput Screening of Neurofilament Light Chain Self-Assembling Peptide for Noninvasive Monitoring of Alzheimer’s Disease

Alzheimer's disease (AD) starts decades before cognitive symptoms develop. Easily accessible and cost-effective biomarkers that accurately reflect AD pathology are essential for both monitoring and therapeutics of AD. Neurofilament light chain (NfL) levels in blood and cerebrospinal fluid are increased in AD more than a decade before the expected onset, thus providing one of the most promising blood biomarkers for monitoring of AD. The clinical practice of employing single-molecule array (Simoa) technology for routine use in patient care is limited by the high costs. Herein, we developed a microarray chip-based high-throughput screening method and screened an attractive self-assembling peptide targeting NfL. Through directly "imprinting" and further analyzing the sequences, morphology, and affinity of the identified self-assembling peptides, the Pep-NfL peptide nanosheet with high binding affinity toward NfL (KD = 1.39 × 10–9 mol/L), high specificity, and low cost was characterized. The superior binding ability of Pep-NfL was confirmed in AD mouse models and cell lines. In the clinical setting, the Pep-NfL peptide nanosheets hold great potential for discriminating between patients with AD (P < 0.001, n = 37), mild cognitive impairment (P < 0.05, n = 26), and control groups (n = 30). This work provides a high-throughput, high-sensitivity, and economical system for noninvasive tracking of AD to monitor neurodegeneration at different stages of disease. The obtained Pep-NfL peptide nanosheet may be useful for assessing dynamic changes in plasma NfL concentrations to evaluate disease-modifying therapies as a surrogate end point of neurodegeneration in clinical trials.