Analytical and Clinical Validity of a High-Throughput, Fully Automated Immunoassay for Plasma Neurofilament Light Chain.

BACKGROUND

Neurofilament light chain (NfL) has emerged as a promising blood-based biomarker for neuroaxonal damage across neurological disorders. Clinical utility has been correlated with disease activity, treatment response, and prognosis. However, routine clinical laboratory implementation requires comprehensive analytical validation to ensure reliability and accuracy.

METHODS

A fully automated Access NfL research use only (RUO) immunoassay was evaluated on the DxI 9000 Immunoassay Analyzer from Beckman Coulter. Analytical validation in plasma assessed precision, analytical sensitivity, linearity, analytical specificity (interference), calibrator and sample stability, and method comparison to the Quanterix Simoa NfL assay. Clinical validity included comparisons of Alzheimer disease (Ad, N = 20) in plasma and multiple sclerosis (MS, N = 16) in serum, vs healthy controls (N = 20 and N = 23, respectively). Plasma-serum equivalence was validated.

RESULTS

The NfL assay met analytical criteria, with precision demonstrating coefficients of variation <3% across runs, a lower limit of quantification of 3.10 pg/mL, linearity with <12% deviation, and <10% interference. Plasma NfL samples remained stable for 18 h at room temperature (<7% difference) and refrigerated (<5%), and through 5 freeze-thaw cycles (<8%). Method comparison yielded high correlation (R2 = 0.981) with positive systematic proportional bias (Passing-Bablok slope = 3.02). Ad patients showed significantly elevated NfL levels (median 49.3 pg/mL, IQR: 28.9-67.8) vs controls (median 29.1 pg/mL, IQR: 22.6-33.8; P = 0.02). MS patients similarly demonstrated higher NfL (median 27.7 pg/mL, IQR: 22.5-50.6) vs controls (median 19.2 pg/mL, IQR: 15.1-25.6; P = 0.003).

CONCLUSIONS

The high-throughput Access NfL assay demonstrated analytical and clinical validity, supporting implementation in research and clinical laboratories.