High throughput detection of the spike glycoprotein of the virus causing COVID-19 by laser diode thermal desorption-triple quadrupole mass spectrometry

激光二极管热解吸-三重四极杆质谱法高通量检测引起 COVID-19 的病毒的刺突糖蛋白

• 批准号: 554601-2020
• 负责人: Segura, PedroAlejandro
• 金额: $ 3.64万
• 依托单位: Université de Sherbrooke
• 依托单位国家: 加拿大
• 项目类别: Alliance Grants
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=705693
• 关键词: throughput; detection; spike; glycoprotein; virus

In order to better manage the current COVID-19 pandemic and support research on the disease in Canada and the rest of the world, it is indispensable to have rapid and sensitive methods of detection of the SARS-CoV-2 virus. Presently, two major types of tests are used: antibody-based and RNA-based. However, in spite of the sensitivity and specificity of these tests, they have important disadvantages such as false negatives in the early stages of infection. The aim of the present proposal is to develop and evaluate the feasibility of a new COVID-19 diagnostic test based on the ultra-fast detection of specific peptides of the spike glycoprotein of the SARS-CoV-2 virus. This protein mediates the entry of virus to target cells and it is one of the pathogen's signature components. The specific peptides of the virus (already identified by another research group) are obtained after proteolytic digestion with trypsin of the spike glycoprotein and they are detected using a high throughput technique, laser diode thermal desorption (LDTD) coupled to triple quadrupole mass spectrometry. If successful, this method could analyze more than 15000 samples in less than 35 hours. The partner organization in this project, Phytronix Technologies, developed the LDTD ionization source and will work closely with academic collaborators to build the method by sharing expertise and resources. The project is divided in four stages necessary to determine the applicability of the method for COVID-19 diagnostic: optimization of instrumental parameters, signal enhancement of the target peptides, extraction and digestion of spike glycoprotein in real conditions and determination of figures of merit of the method (limits of detection, precision, rate of false positives and false negatives, etc.). Phytronix Technologies will facilitate the deployment of the method by developing turnkey solutions for potential customers. These turnkey solutions (simplified protocols, reagent kits) allow the proper transfer of analytical methods to the customers by allowing staff with minimal training to use the developed method. Such effort is a key step for knowledge transfer to users and a critical step towards facilitating the diffusion of the results of the present project.

为了更好地管理当前的新冠肺炎疫情，并支持加拿大和世界其他地区对该疾病的研究，拥有快速而灵敏的检测SARS-CoV-2病毒的方法是必不可少的。目前，主要使用两种类型的检测：基于抗体的检测和基于RNA的检测。然而，尽管这些测试具有敏感性和特异性，但它们也有重要的缺点，如在感染的早期阶段出现假阴性。本建议的目的是开发和评估一种新的新冠肺炎诊断试验的可行性，该试验基于对SARS-CoV-2病毒刺突糖蛋白特异性多肽的超快速检测。这种蛋白质介导病毒进入靶细胞，是病原体的标志性成分之一。病毒的特定多肽(已由另一个研究小组鉴定)是在用胰酶消化尖峰糖蛋白后获得的，并使用高通量技术-激光二极管热解吸(LDTD)-三重四极杆质谱仪进行检测。如果成功，这种方法可以在不到35小时内分析15000多个样本。该项目的合作伙伴组织Phytronix Technologies开发了LDTD电离源，并将与学术合作者密切合作，通过共享专业知识和资源来构建该方法。该项目分为四个阶段，以确定该方法对新冠肺炎诊断的适用性：仪器参数的优化，目标多肽的信号增强，真实条件下尖峰糖蛋白的提取和消化，以及方法优值系数的确定(检测限、精密度、假阳性和假阴性率等)。Phytronix技术公司将通过为潜在客户开发交钥匙解决方案来促进该方法的部署。这些交钥匙解决方案(简化的规程、试剂盒)允许经过最少培训的工作人员使用开发的方法，从而将分析方法适当地传递给客户。这种努力是向用户转让知识的关键一步，也是促进传播本项目成果的关键一步。