Objective: To evaluate CSF α-syn-SAA interlaboratory reproducibility in diagnosing Parkinson´s Disease (PD) and atypical parkinsonisms as well as interrogate the effect of changes in lot of α-syn recombinant in assay kinetics.

Background: Seed amplification assays (SAAs) demonstrate exceptional accuracy in diagnosing synucleinopathies like PD and multiple system atrophy (MSA) distinguishing them from other atypical parkinsonian disorders such as progressive supranuclear palsy (PSP). However, the lack of universally accepted standards, along with variations in reagents, protocols, and instrumentation, raises concerns about interlaboratory reproducibility and hinders widespread clinical translation. In this study, we assess the CSF α-syn-SAA reproducibility across two different labs with independent equipment and personnel both at Hospital Clinic and IDIBAPS. We evaluated the consistency of positive and negative outcomes as well as differences in assay kinetics between labs and lot-to-lot α-syn recombinant.

Method: Sporadic PD, MSA and PSP CSF samples were analyzed independently in two specialized laboratories: Lab1 at Hospital Clínic (Spain) and Lab2 at IDIBAPS (Spain). Both laboratories developed and standardized their CSF αSyn-SAA protocols independently. All sample sets were tested in a blinded manner with results reported as positive/negative. In addition, two different lots of protein recombinant were tested at Lab2 for lot effect in consistency and assay kinetics

Results: Upon comparison, we found an 80% agreement and a Cohen’s Kappa of 0.364, indicating reproducibility of CSF α-syn-SAA between laboratories. Changes in recombinant lots did not affect final outcomes, maintaining an overall 80% sensitivity for disease diagnosis and yielding a 93% agreement and 0.445 Cohen’s Kappa. However, switching recombinant lots under the same protocol settings introduced variability in time to threshold, and time to 50% fluorescence.

Conclusion: Our results demonstrate that CSF α-syn-SAA is a reliable and reproducible method for identifying PD and distinguishing it from PSP across different laboratories with high sensitivity and specificity. While variations in recombinant lot may influence reaction kinetics, they do not impact the final diagnosis. This validation is a crucial step toward establishing SAA as a dependable diagnostic tool for PD in clinical practice.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/interlaboratory-cross-validation-of-csf-%ce%b1-synuclein-seeding-aggregation-assay-for-parkinsonisms/