(T1030-02-06) Qualification of the S-PLEX Neurology Panel 1 (GFAP, NfL and Tau) MSD Assay

Purpose: Neurofilament light chain (NfL), glial fibrillary acidic protein (GFAP) and Tau have been identified as valuable diagnostic and prognostic biomarkers of brain injury and other neurological disorders¹. In the context of a clinical trial, data from such measurements may play a critical role in informing treatment efficacy and safety evaluations, patient enrollments as well as stratification. Assays which evaluate these analytes in plasma and CSF can be used to measure changes in each matrix in response to treatment. Ultimately his data can be used to support assessment of the biomarkers in plasma, which is a much less invasive sample to collect compared to CSF, in later phase clinical trials. Leveraging multiplexed panels for these biomarkers enables multiple analytes to be measured simultaneously from a single sample, which is especially advantageous for testing rare or volume limited samples. Platforms such as MSD offer a wide dynamic quantitation range and sensitivity with the added benefit of low sample volumes unlike conventional platforms such as ELISA. Here, we describe the qualification of an S-PLEX Neurology Panel 1 (NfL, Tau and GFAP) in, both, plasma and CSF matrices.

Methods: The Neurology Panel 1 was procured from Meso Scale Diagnostics (MSD). Plasma and CSF from normal and Parkinson’s Disease individuals were obtained from BioIVT. The assay was performed per manufacturer’s instructions and qualified for Accuracy & Precision, Parallelism, Stability, and Ruggedness. Ten normal donors were first evaluated to determine the endogenous levels of each of the 3 analytes in both plasma and CSF, and their placement on the standard curve, respectively. This not only aided in selecting the appropriate starting sample dilution factor for each of the analytes, but also in the preparation of 5 levels of endogenous qualification samples (ULOQ, HQC, MQC, LQC and LLOQ) in both matrices.

Results: The S-PLEX method for the Neurology Panel 1 was qualified in plasma with the quantitation range from 479442 – 3016 fg/mL for GFAP, 5495822- 2469 fg/mL for NfL and 225775- 242 fg/mL for Tau with an MRD of 1:2. Similarly, the panel was qualified in CSF with the quantitation range from 5358116– 1822 fg/mL for GFAP, 31065114- 95316 fg/mL for NfL and 1134236- 3952 fg/mL for Tau with an MRD of 1:10. Both matrices across all 3 analytes demonstrated acceptable Accuracy & Precision, Parallelism, Short-term and Freeze-Thaw Stability, and Ruggedness as well as a lack of Prozone effect, except for GFAP, which failed 24 hr RT stability.

Conclusion: The S-PLEX Neurology Panel 1 was successfully qualified in CSF and plasma. The qualified method would be applied in a clinical study to help explore the endogenous concentrations of GFAP, NfL and Tau in both the matrices. This assay combines 3 essential analytes for multiple neurological indications using the novel S-Plex methodology, demonstrating precision and accuracy to low fg/mL concentrations, which is a critical concern for these neural biomarkers where sample volume is extremely limited.

References: 1. Park Y, Kc N, Paneque A, Cole PD. Tau, Glial Fibrillary Acidic Protein, and Neurofilament Light Chain as Brain Protein Biomarkers in Cerebrospinal Fluid and Blood for Diagnosis of Neurobiological Diseases. Int J Mol Sci. 2024 Jun 7;25(12):6295.