# Analytical validation of a biochemical test for alpha-synuclein aggregates in biological fluids for the diagnosis of Parkinson's Disease

> **NIH NIH U44** · AMPRION, INC. · 2021 · $809,593

## Abstract

PROJECT SUMMARY
 Misfolded protein aggregates are toxic supramolecular proteinaceous conglomerates that self-
replicate in the host, causing damage to surrounding tissue. Several of these protein aggregates are
strongly associated with neurodegenerative diseases, such as the prion protein in prion diseases,
Amyloid β and tau in Alzheimer’s (AD), TDP-34 in amyotrophic lateral sclerosis (ALS) and
Frontotemporal dementia (FTD), and α-synuclein (αS) in Parkinson’s disease (PD), dementia with
Lewy bodies (DLB) and Multiple Systemic Atrophy (MSA). The molecular mechanisms behind the in
vivo formation of these aggregates and their effect on neural tissue remain elusive, but their
compelling implication as early drivers of the disease processes identifies them as potentially useful
disease biomarkers and therapeutic targets. The primary goal of this proposal is to perform robust
analytical validation of a Protein Misfolding Cyclic Amplification (PMCA) assay for detection of αS
aggregates in cerebrospinal fluid (CSF) and plasma of patients with synucleinopathies. The PMCA
platform uses the intrinsic self-replicating nature of misfolded protein aggregates to cyclically amplify
minute quantities of αS aggregates in biological samples. The assay is performed in a 96-well plate,
and the amplification is monitored over time using the fluorescent amyloid-specific dye, ThioflavinT.
Method development is focused on robustness to support future use in a controlled laboratory with
high sample throughput and strict requirements for assay accuracy and reproducibility. Analytical
validation will be performed in accordance with FDA and Clinical Laboratory Standard Institute (CLSI)
recommendations, using a combination of clinically positive patient samples and samples spiked with
synthetically generated aggregates to evaluate method accuracy, precision, sensitivity, specificity,
reportable range, and other critical assay characteristics. The overall objectives for phase I are to
finalize standard operating procedures and protocols and perform analytical validation of
qualitative/semi-quantitative αS-PMCA assays for detection of αS oligomers in CSF and plasma for
use as a diagnostic biomarker for PD and related synucleinopathies. The overall objectives for phase
II are to adapt the assays for quantitation/kinetic characterization of αS oligomers in CSF and plasma,
and to explore expanded context of use to include indications as: a susceptibility/risk biomarker to
identify patients at risk of developing synucleinopathies prior to onset of clinical symptoms; a
diagnostic biomarker to distinguish between different type of synucleinopathies; a monitoring
biomarker to assess disease progression or treatment effectiveness; and a prognostic biomarker with
ability to characterize disease type and severity.

## Key facts

- **NIH application ID:** 10361903
- **Project number:** 4U44NS111672-02
- **Recipient organization:** AMPRION, INC.
- **Principal Investigator:** Karen MacLeod
- **Activity code:** U44 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $809,593
- **Award type:** 4N
- **Project period:** 2021-05-01 → 2023-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10361903

## Citation

> US National Institutes of Health, RePORTER application 10361903, Analytical validation of a biochemical test for alpha-synuclein aggregates in biological fluids for the diagnosis of Parkinson's Disease (4U44NS111672-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10361903. Licensed CC0.

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