Streamlined alpha-synuclein RT-QuIC assay for various biospecimens in Parkinson's disease and dementia with Lewy bodies.
Streamlined alpha-synuclein RT-QuIC assay for various biospecimens in Parkinson's disease and dementia with Lewy bodies.
复制标题
DOI：
10.1186/s40478-021-01175-w
复制
发表时间：
2021-04-07
影响因子：
7.1
通讯作者：
Chen SG
中科院分区：
文献类型：
--
作者：
Bargar C;Wang W;Gunzler SA;LeFevre A;Wang Z;Lerner AJ;Singh N;Tatsuoka C;Appleby B;Zhu X;Xu R;Haroutunian V;Zou WQ;Ma J;Chen SG
帕金森氏病(PD)和路易体痴呆(DLb)的确诊依赖于尸检发现的疾病相关α-突触核蛋白(α-Synd)是中枢神经系统(CNS)中错误折叠的蛋白质聚集体。用于超灵敏检测αSynd聚集体的实时抖动诱导转换(RT-QuIC)方法的最新进展使临床上可获得的生物标本(包括脑脊液和周围组织)的诊断价值重新焕发生机。然而，目前的αSYN RT-QIC分析平台差异很大，因此在广泛的生物标本和不同的实验室中实施和标准化αSYD的测量是具有挑战性的。我们已经简化了基于完全由商业试剂组装的第二代检测平台的αSyn RT-QuIC检测。简化的RT-QuIC方法包括一种简化的方法，需要最少的实际操作时间，并允许统一分析来自PD和DLB的不同类型的生物检材中的αSynd。在百万倍稀释的脑组织匀浆和PD和DLB病例的脑脊液纳米升中实现了对αSynd聚集体的超灵敏和特异的RT-QuIC检测，但不能从对照中检测到。比较分析表明，在脑组织匀浆和脑脊液中，αSynd在DLb中的播种活性均高于PD。我们的方法进一步验证了来自组织仓库的214例神经病理确诊病例的脑脊液样本(88例PD、58例DLB和68例对照)，其敏感性为98%，特异性为100%。最后，采用单一的RT-QuIC分析方法，统一检测帕金森病患者不同类型组织中α-Synd的种子活性，这些组织包括大脑、皮肤、唾液腺和结肠。我们预计，我们简化的方案将使感兴趣的实验室能够轻松、快速地对PD和DLB的各种临床标本实施αSyn RT-QuIC检测。所有检测成分的商业化产品的使用将提高RT-QuIC检测的稳健性和标准化，用于不同地点的诊断应用。由于样品消耗量极低，这种超灵敏的RT-QuIC检测方法将有助于有效利用和共享稀缺的生物检疫菌资源。我们简化的RT-QIC方法适合于追踪α综合征在中枢神经系统和患者外周组织中的分布。正在进行的αSynd RT-QuIC分析作为临床可获得的生物样本中PD和DLB的潜在生物标志物的评估，对于了解疾病的发病机制，改进早期和鉴别诊断，以及在临床试验中监测治疗效果具有广泛的意义。
Definitive diagnosis of Parkinson’s disease (PD) and dementia with Lewy bodies (DLB) relies on postmortem finding of disease-associated alpha-synuclein (αSynD) as misfolded protein aggregates in the central nervous system (CNS). The recent development of the real-time quaking induced conversion (RT-QuIC) assay for ultrasensitive detection of αSynD aggregates has revitalized the diagnostic values of clinically accessible biospecimens, including cerebrospinal fluid (CSF) and peripheral tissues. However, the current αSyn RT-QuIC assay platforms vary widely and are thus challenging to implement and standardize the measurements of αSynD across a wide range of biospecimens and in different laboratories. We have streamlined αSyn RT-QuIC assay based on a second generation assay platform that was assembled entirely with commercial reagents. The streamlined RT-QuIC method consisted of a simplified protocol requiring minimal hands-on time, and allowing for a uniform analysis of αSynD in different types of biospecimens from PD and DLB. Ultrasensitive and specific RT-QuIC detection of αSynD aggregates was achieved in million-fold diluted brain homogenates and in nanoliters of CSF from PD and DLB cases but not from controls. Comparative analysis revealed higher seeding activity of αSynD in DLB than PD in both brain homogenates and CSF. Our assay was further validated with CSF samples of 214 neuropathologically confirmed cases from tissue repositories (88 PD, 58 DLB, and 68 controls), yielding a sensitivity of 98% and a specificity of 100%. Finally, a single RT-QuIC assay protocol was employed uniformly to detect seeding activity of αSynD in PD samples across different types of tissues including the brain, skin, salivary gland, and colon. We anticipate that our streamlined protocol will enable interested laboratories to easily and rapidly implement the αSyn RT-QuIC assay for various clinical specimens from PD and DLB. The utilization of commercial products for all assay components will improve the robustness and standardization of the RT-QuIC assay for diagnostic applications across different sites. Due to ultralow sample consumption, the ultrasensitive RT-QuIC assay will facilitate efficient use and sharing of scarce resources of biospecimens. Our streamlined RT-QuIC assay is suitable to track the distribution of αSynD in CNS and peripheral tissues of affected patients. The ongoing evaluation of RT-QuIC assay of αSynD as a potential biomarker for PD and DLB in clinically accessible biospecimens has broad implications for understanding disease pathogenesis, improving early and differential diagnosis, and monitoring therapeutic efficacies in clinical trials.
登录
查看更多内容
影响因子：
4.6
作者：
Hoover, Clare E.;Davenport, Kristen A.;Hoover, Edward A.
通讯作者：
Hoover, Edward A.
影响因子：
12.7
作者：
Doppler K;Ebert S;Uçeyler N;Trenkwalder C;Ebentheuer J;Volkmann J;Sommer C
通讯作者：
Sommer C
影响因子：
4.1
作者：
Kalia, Lorraine V.
通讯作者：
Kalia, Lorraine V.
影响因子：
8.6
作者：
Kang, Un Jung;Boehme, Amelia K.;Soto, Claudio
通讯作者：
Soto, Claudio
DOI：
10.1097/paf.0000000000000420
发表时间：
2018-12-01
影响因子：
1
作者：
Garland, Jack;Philcox, Winston;Tse, Rexson
通讯作者：
Tse, Rexson