Session: Session 5
Characterisation of candidate primary calibrator materials for P-Tau measurements in biological fluids
Background
NEuroBioStand is an EU-funded project aimed at developing a metrological research framework for standardising blood-based biomarkers of neurodegenerative diseases (NDDs) with the objective of implementation and commercialisation of promising assays for NDD biomarkers fulfilling requirements of the in vitro diagnostic regulation (IVDR). P-tau is currently included in the AT(N) framework for AD diagnosis together with other biomarkers and amyloid-β and tau in CSF have been developed into regulatory approved biomarkers in CSF. The standardisation of the measurements for this biomarker is important to establish common cut-off values and reference ranges.
Methods
To achieve this, recombinant phosphorylated proteins and synthetic phosphopeptides were sourced as candidate primary calibrators. These materials underwent rigorous evaluation for purity and mass fraction using liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) and amino acid analysis (AAA).
Results
The first step towards the standardisation of P-tau is the definition of the measurand. Three-phosphorylated isoforms, namely P-tau181, P-tau217, and P-tau231, were prioritized with the objective of developing fit-for-purpose calibrators characterized for purity and mass fraction. Additionally, a SI-traceable reference measurement procedure (RMP) in plasma is under development. This procedure aims to standardize concentrations of phosphorylated epitopes, enabling the calculation of phosphorylated to non-phosphorylated fragment ratios.
Conclusions
NEuroBioStand consortium gathers national metrology institutes, clinicians, academics and
IVD-providers to achieve traceability to SI units for fluid biomarkers in NDD. The development
of a RMP will allow the certification of candidate matrix-based reference materials
that will be used to harmonise measurements of P-tau in plasma across different analytical platforms.
Methods
To achieve this, recombinant phosphorylated proteins and synthetic phosphopeptides were sourced as candidate primary calibrators. These materials underwent rigorous evaluation for purity and mass fraction using liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) and amino acid analysis (AAA).
Results
The first step towards the standardisation of P-tau is the definition of the measurand. Three-phosphorylated isoforms, namely P-tau181, P-tau217, and P-tau231, were prioritized with the objective of developing fit-for-purpose calibrators characterized for purity and mass fraction. Additionally, a SI-traceable reference measurement procedure (RMP) in plasma is under development. This procedure aims to standardize concentrations of phosphorylated epitopes, enabling the calculation of phosphorylated to non-phosphorylated fragment ratios.
Conclusions
NEuroBioStand consortium gathers national metrology institutes, clinicians, academics and
IVD-providers to achieve traceability to SI units for fluid biomarkers in NDD. The development
of a RMP will allow the certification of candidate matrix-based reference materials
that will be used to harmonise measurements of P-tau in plasma across different analytical platforms.