Search Results (8)

Introduction and objective: Assessment of α-synuclein (αSyn) seed amplification assays (αSyn-SAA) accuracy in distinguishing Parkinson’s disease (PD) from controls using cerebrospinal fluid (CSF), blood, skin, extracellular vesicles (ECV), saliva, olfactory mucosa (OM), gastrointestinal tract (GIT), and submandibular gland (SMG). Methodology: PubMed was searched for articles from 2010 to January 2025. The quality assessment used robvis. Diagnostic values with a 95% confidence interval (CI) were obtained. Z-test, Wald CI, and ANOVA were performed. Diagnostic odds ratio (DOR) was used. Results: αSyn-SAAs showed strong diagnostic performance in distinguishing PD from controls across various tissue and fluid types. Overall, αSyn-SAAs demonstrated high sensitivity (86%) and specificity (92%). Among all biomatrices, CSF, skin, blood, and ECV yielded the highest diagnostic accuracy, with sensitivity and specificity approaching or exceeding 90%. In contrast, saliva, oral mucosa, and gastrointestinal tract samples showed more modest sensitivity, though specificity remained relatively high. ECV, CSF, skin, and blood matrices also demonstrated the highest DOR, supporting their potential clinical utility. Conclusions: ECV and blood warrant priority in αSyn-SAA for high accuracy and minimal invasiveness, while GIT, OM, and oral samples show limited utility; saliva and SMG need refinement. Full article

Lewy Body Disease (LBD) and Multiple System Atrophy (MSA) are synucleinopathies with distinct prognoses and neuropathologies, however, with overlapping clinical symptoms. Different disease characteristics are proposed to be determined by distinct conformations of alpha-synuclein (α-Syn) aggregates, which can self-propagate and spread between cells via a prion-like mechanism. The goal of this study is to investigate whether α-syn aggregates amplified from brain and CSF samples of LBD and MSA patients using the Seed Amplification Assay (SAA) maintain α-Syn seeding properties similar to those of α-syn aggregates derived from patients’ brains. To address this, SAA-amplified and un-amplified α-Syn aggregates from LBD and MSA patients’ brains, as well as SAA-amplified α-Syn aggregates from LBD and MSA patients’ CSF samples, were used to treat synuclein biosensor cells, and induced intracellular α-Syn inclusions were analyzed by confocal microscopy. Our data indicate that induced α-Syn aggregates from LBD and MSA patients’ brains have similar seeding properties and morphological characteristics in the α-Syn biosensor cells as those amplified from LBD and MSA patients’ brains, as well as those amplified from LBD and MSA patients’ CSF samples. In this study, we demonstrated that, regardless of the source of aggregates, the seeds from LBD and MSA produce cellular accumulation of α-Syn with distinct morphologies, confirming the presence of different conformational strains of α-Syn in LBD and MSA and allowing us to differentiate synucleinopathies based on the morphology of aggregates and seeding properties. Full article

Parkinson’s disease (PD) is the second most common neurodegenerative disease in the world. Currently, PD is incurable, and the diagnosis of PD mainly relies on clinical manifestations. The central pathological event in PD is the abnormal aggregation and deposition of misfolded α-synuclein (α-Syn) protein aggregates in the Lewy body (LB) in affected brain areas. Behaving as a prion-like seeding, the misfolded α-syn protein can induce and facilitate the aggregation of native unfolded α-Syn protein to aggravate α-Syn protein aggregation, leading to PD progression. Recently, in a blood-based α-Syn seeding amplification assay (SAA), Kluge et al. identified pathological α-Syn seeding activity in PD patients with Parkin (PRKN) gene variants. Additionally, pathological α-syn seeding activity was also identified in sporadic PD and PD patients with Leucine-rich repeat kinase 2 (LRRK2) or glucocerebrosidase (GBA) gene variants. Principally, the α-Syn SAA can be used to detect pathological α-Syn seeding activity, which will significantly enhance PD diagnosis, progression monitoring, prognosis prediction, and anti-PD therapy. The significance and future strategies of α-Syn SAA protocol are highlighted and proposed, whereas challenges and limitations of the assay are discussed. Full article

The present investigation aims to examine the role of α-synuclein seed amplification assays for screening Parkinson’s disease. Parkinson’s disease (PD) is a debilitating neurodegenerative disorder caused by the loss of dopaminergic neurons in the midbrain, leading to symptoms such as tremors, bradykinesia, postural instability, dementia, and depression. It is classified as an α-synucleinopathy related to the role of α-synuclein aggregates in neuron degeneration. Diagnosis relies on clinical assessment without premortem diagnostic tests or imaging, often resulting in delayed detection and impaired symptom management. In this regard, our study explores a screening technique using an amplification assay to measure α-synuclein levels in cerebrospinal fluid, which could potentially identify early pathological changes and improve diagnostic accuracy and patient outcomes. While preliminary results are promising, further studies are needed to evaluate this approach’s accuracy and clinical feasibility. A review of numerous trials demonstrates that α-synuclein seeding amplification assays (SAA) are a highly reliable, sensitive, and specific diagnostic tool for PD. This assay offers a promising opportunity to improve early diagnosis and quantify severity, especially for asymptomatic individuals or those with a family history of PD, allowing for earlier intervention and more effective disease management. In summary, the emerging body of evidence supporting α-synuclein as a biomarker should allow patients with PD to be detected and treated sooner, enhancing patients’ quality of life and potentially changing the disease trajectory. Full article

Multiple system atrophy and Lewy body diseases (LBDs) such as Parkinson’s disease, dementia with Lewy bodies, and Parkinson’s disease with dementia, known as synucleinopathies, are defined neuropathologically by the accumulation and deposition of aberrant protein aggregates, primarily in neuronal cells. Seeding aggregation assays (SAA) have significant potential as biomarkers for early diagnosis, monitoring disease progression, and evaluating treatment efficacy for these diseases. Real-time quaking-induced conversion (RT-QuIC) and Protein Misfolding Cyclic Amplification (PMCA) assays represent two ultrasensitive protein amplification techniques that were initially tested for the field of prion disorders. Although the fundamental idea behind the creation of these two methods is very similar, their technical differences resulted in different levels of diagnostic accuracy for the identification of prion proteins, making the RT-QuIC assay the most trustworthy and effective instrument for the detection of suspected cases of LBDs and prion-like diseases. Full article

Alpha-synuclein seed amplification assays (αSyn-SAAs) have emerged as promising diagnostic tools for Parkinson’s disease (PD) by detecting misfolded αSyn and amplifying the signal through cyclic shaking and resting in vitro. Recently, our group and others have shown that multiple biospecimens, including CSF, skin, and submandibular glands (SMGs), can be used to seed the aggregation reaction and robustly distinguish between patients with PD and non-disease controls. The ultrasensitivity of the assay affords the ability to detect minute quantities of αSyn in peripheral tissues, but it also produces various technical challenges of variability. To address the problem of variability, we present a high-yield αSyn protein purification protocol for the efficient production of monomers with a low propensity for self-aggregation. We expressed wild-type αSyn in BL21 Escherichia coli, lysed the cells using osmotic shock, and isolated αSyn using acid precipitation and fast protein liquid chromatography (FPLC). Following purification, we optimized the ionic strength of the reaction buffer to distinguish the fluorescence maximum (Fmax) separation between disease and healthy control tissues for enhanced assay performance. Our protein purification protocol yielded high quantities of αSyn (average: 68.7 mg/mL per 1 L of culture) and showed highly precise and robust αSyn-SAA results using brain, skin, and SMGs with inter-lab validation. Full article

Recent studies have been able to detect α-synuclein (αSyn) seeding in formaldehyde-fixed paraffin-embedded (FFPE) tissues from patients with synucleinopathies using seed amplification assays (SAAs), but with relatively low sensitivity due to limited protein extraction efficiency. With the aim of introducing an alternative option to frozen tissues, we developed a streamlined protein extraction protocol for evaluating disease-specific seeding in FFPE human brain. We evaluated the protein extraction efficiency of different tissue preparations, deparaffinizations, and protein extraction buffers using formaldehyde-fixed and FFPE tissue of a single Lewy body disease (LBD) subject. Alternatively, we incorporated heat-induced antigen retrieval and dissociation using a commercially available kit. Our novel protein extraction protocol has been optimized to work with 10 sections of 4.5-µm-thickness or 2-mm-diameter micro-punch of FFPE tissue that can be used to seed SAAs. We demonstrated that extracted proteins from FFPE still preserve seeding potential and further show disease-specific seeding in LBD and multiple system atrophy. To the best of our knowledge, our study is the first to recapitulate disease-specific αSyn seeding behaviour in FFPE human brain. Our findings open new perspectives in re-evaluating archived human brain tissue, extending the disease-specific seeding assays to larger cohorts to facilitate molecular subtyping of synucleinopathies. Full article

Various disease-associated forms or strains of α-synuclein (αSyn^D) can spread and accumulate in a prion-like fashion during synucleinopathies such as Parkinson’s disease (PD), Lewy body dementia (DLB), and multiple system atrophy (MSA). This capacity for self-propagation has enabled the development of seed amplification assays (SAAs) that can detect αSyn^D in clinical samples. Notably, α-synuclein real-time quaking-induced conversion (RT-QuIC) and protein misfolding cyclic amplification (PMCA) assays have evolved as ultrasensitive, specific, and relatively practical methods for detecting αSyn^D in a variety of biospecimens including brain tissue, CSF, skin, and olfactory mucosa from synucleinopathy patients. However, αSyn SAAs still lack concordance in detecting MSA and familial forms of PD/DLB, and the assay parameters show poor correlations with various clinical measures. End-point dilution analysis in αSyn RT-QuIC assays allows for the quantitation of relative amounts of αSyn^D seeding activity that may correlate moderately with clinical measures and levels of other biomarkers. Herein, we review recent advancements in α-synuclein SAAs for detecting αSyn^D and describe in detail the modified Spearman–Karber quantification algorithm used with end-point dilutions. Full article