变异型克雅氏病朊病毒感染的检测：一种基于血液的检测方法。

Detection of prion infection in variant Creutzfeldt-Jakob disease: a blood-based assay.

机构信息

MRC Prion Unit, Department of Neurodegenerative Disease, UCL Institute of Neurology, London, UK.

出版信息

Lancet. 2011 Feb 5;377(9764):487-93. doi: 10.1016/S0140-6736(10)62308-2.

DOI:10.1016/S0140-6736(10)62308-2

PMID:21295339

Abstract

BACKGROUND

Variant Creutzfeldt-Jakob disease (vCJD) is a fatal neurodegenerative disorder originating from exposure to bovine-spongiform-encephalopathy-like prions. Prion infections are associated with long and clinically silent incubations. The number of asymptomatic individuals with vCJD prion infection is unknown, posing risk to others via blood transfusion, blood products, organ or tissue grafts, and contaminated medical instruments. We aimed to establish the sensitivity and specificity of a blood-based assay for detection of vCJD prion infection.

METHODS

We developed a solid-state binding matrix to capture and concentrate disease-associated prion proteins and coupled this method to direct immunodetection of surface-bound material. Quantitative assay sensitivity was assessed with a serial dilution series of 10⁻⁷ to 10⁻¹⁰ of vCJD prion-infected brain homogenate into whole human blood, with a baseline control of normal human brain homogenate in whole blood (10⁻⁶). To establish the sensitivity and specificity of the assay for detection of endogenous vCJD, we analysed a masked panel of 190 whole blood samples from 21 patients with vCJD, 27 with sporadic CJD, 42 with other neurological diseases, and 100 normal controls. Samples were masked and numbered by individuals independent of the assay and analysis. Each sample was tested twice in independent assay runs; only samples that were reactive in both runs were scored as positive overall.

FINDINGS

We were able to distinguish a 10⁻¹⁰ dilution of exogenous vCJD prion-infected brain from a 10⁻⁶ dilution of normal brain (mean chemiluminescent signal, 1·3×10⁵ [SD 1·1×10⁴] for vCJD vs 9·9×10⁴ [4·5×10³] for normal brain; p<0·0001)—an assay sensitivity that was orders of magnitude higher than any previously reported. 15 samples in the masked panel were scored as positive. All 15 samples were from patients with vCJD, showing an assay sensitivity for vCJD of 71·4% (95% CI 47·8–88·7) and a specificity of 100% (95% CIs between 97·8% and 100%).

INTERPRETATION

These initial studies provide a prototype blood test for diagnosis of vCJD in symptomatic individuals, which could allow development of large-scale screening tests for asymptomatic vCJD prion infection.

FUNDING

UK Medical Research Council.

摘要

背景

变异型克雅氏病（vCJD）是一种致命的神经退行性疾病，源于接触牛海绵状脑病样朊病毒。朊病毒感染与漫长且临床无症状的潜伏期有关。vCJD 朊病毒感染的无症状个体数量未知，通过输血、血液制品、器官或组织移植物以及受污染的医疗器械，对他人构成风险。我们旨在建立一种基于血液的检测方法，用于检测 vCJD 朊病毒感染。

方法

我们开发了一种固态结合基质来捕获和浓缩与疾病相关的朊病毒蛋白，并将这种方法与表面结合物质的直接免疫检测相结合。通过将 10⁻⁷ 至 10⁻¹⁰ 的 vCJD 朊病毒感染性脑匀浆系列稀释到全血中，评估定量检测的敏感性，以全血中正常人脑匀浆作为基线对照（10⁻⁶）。为了确定该检测方法用于检测内源性 vCJD 的敏感性和特异性，我们分析了一个由 21 名 vCJD 患者、27 名散发性克雅氏病患者、42 名其他神经疾病患者和 100 名正常对照者的 190 份全血样本组成的盲样组。样本由独立于检测和分析的个体进行编号和标记。每个样本在两次独立的检测运行中进行检测；只有在两次运行中均呈阳性的样本才被整体判定为阳性。

结果

我们能够将 10⁻¹⁰ 稀释的外源性 vCJD 朊病毒感染性脑匀浆与 10⁻⁶ 稀释的正常脑匀浆区分开来（vCJD 的平均化学发光信号为 1.3×10⁵[标准差为 1.1×10⁴]，正常脑匀浆为 9.9×10⁴[4.5×10³]；p<0.0001），检测敏感性比以前报道的高几个数量级。盲样组中的 15 个样本被判定为阳性。所有 15 个样本均来自 vCJD 患者，检测 vCJD 的敏感性为 71.4%（95%置信区间为 47.8%至 88.7%），特异性为 100%（95%置信区间在 97.8%至 100%之间）。