High-throughput proteomics using submicroliter amounts of plasma for comprehensive assessment of the immune status

使用亚微升血浆进行高通量蛋白质组学综合评估免疫状态

基本信息

批准号：
10381719
负责人：
Hanno Steen
金额：
$ 117.82万
依托单位：
BOSTON CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-09 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10381719
关键词：
Adolescent Adult Agreement Aliquot Antibodies Antibody titer measurement Autoimmunity Bacterial Infections Biological Assay Biological Markers Blood Plasma Volume Blood Volume Boston C-reactive protein Cells Communicable Diseases Communities Complement Consumption Data Development Disease Elderly Erythrocyte Sedimentation Rate Fibrinogen Frail Elderly Funding Funding Opportunities Future Health Hepatitis B Surface Antigens Hepatitis B Vaccination Hepatitis B Vaccines Human Human body Hypersensitivity Immune Immune response Immune system Immunity Immunoglobulin Domain Immunoglobulins Immunologic Deficiency Syndromes Immunologic Monitoring Immunology Individual Infection Maps Measures Methods Modality Molecular Monitor Mycoses National Institute of Allergy and Infectious Disease Neonatal Newborn Infant Organism Parasitic infection Participant Patients Pediatric Hospitals Persons Plasma Plasma Proteins Population Populations at Risk Preparation Prevention Proteins Proteome Proteomics Regulation Research Research Design Resolution Sampling Serology Set protein Specimen System Systems Biology Term Birth Testing Time Transplantation Vaccine Design Virus Diseases Vulnerable Populations Work age group anti-hepatitis B base clinical biomarkers cohort complement pathway complement system experimental study gene product immune function immunological status immunoregulation improved innovation insight interest liquid chromatography mass spectrometry milliliter nanolitre novel premature preterm newborn procalcitonin response vaccine immunogenicity vaccinology

项目摘要

SUMMARY Often, many different tests are performed to obtain a multiparameter assessment of the innate and adaptive immune status. Such tests often include erythrocyte sedimentation rate, C-reactive protein (CRP), procalcitonin, various complement pathway proteins, total antibody (Ab) isotypes and/or subclasses, and specific Ab titers. All these tests are performed on different analytical platforms requiring a separate aliquot for each test. Thus, even with modest sample consumption per test, there is a substantial cumulative volume required for multiple independent tests. While these sample requirements may not be prohibitive for healthy adolescents and adults, they are particularly challenging in especially vulnerable populations such as premature newborns, and frail and/or elderly individuals. For these populations, a wide range of different tests is often very important, however, samples are often limited. In agreement with the purpose of the funding opportunity “Development of Sample Sparing Assays for Monitoring Immune Responses”, we propose to move four different protein assays onto a common processing and analysis platform, which can be performed on the same submicroliter aliquot of plasma thereby significantly reducing the sample volume requirements. The four protein assays of interest include i) an expanded panel of complement components, ii) CRP, iii) antibody isotypes and subclasses including a novel and innovative immunoglobulin domain-resolved map of the IgOme-related protein repertoire (aka IgOme), and iv) the classical plasma proteome, i.e. ~400 well described proteins many of which of clear immunological function. The quantitative information from all these assays will provide us with an exquisite map of the immune status, a hypothesis that we will be tested by correlating our findings with the anti-hepatitis B titer measured before and after hepatitis B vaccination in >500 newborns. Samples will be provided by Boston Children's Hospital's NIAID-funded HIPC (Human Immunology Project Consortium), which will significantly increase the impact of this cohort beyond its original scope. Taking the request of this funding opportunity for sample sparing to a different level, we will advance our plasma proteomics pipeline to quadruple the throughput to at least 96 samples/day and to quarter the plasma volume requirements to <100 nanoliter. Thus, in the future, 1 microliter of plasma will be sufficient for tens of temporally independent LC/MS- based protein assays, resulting in quantitative information for hundreds of proteins such that even a miniscule amount of plasma from the smallest and/or most frail patients will be sufficient to establish an exquisitely detailed and unbiased molecular map of immune status of any person.

概括通常，会进行许多不同的测试以获得先天和后天的多参数评估。适应性免疫状态通常包括红细胞沉降率、C反应蛋白。 (CRP)、降钙素原、各种补体途径蛋白、抗体总 (Ab) 同种型和/或所有这些测试都是在不同的分析平台上进行的。每次测试都需要单独的等分试样，因此，即使每次测试的样品消耗量不大，是多个独立测试所需的大量累积体积。对于健康的青少年和成年人来说，这些要求可能不会令人望而却步，但它们尤其是对于早产儿、体弱者和/或老年人等特别脆弱的人群来说具有挑战性然而，对于这些人群来说，广泛的不同测试通常非常重要。样本往往是有限的。与资助机会“开发样品备用测定”的目的一致 “监测免疫反应”，我们建议将四种不同的蛋白质检测转移到一个共同的处理和分析平台，可在相同的亚微升血浆等分试样上进行从而显着减少了四种感兴趣的蛋白质测定的样品体积要求。包括 i) 补体成分的扩展组，ii) CRP，iii) 抗体同种型和亚类包括 IgOme 相关蛋白的新型免疫球蛋白结构域解析图谱库（又名 IgOme），以及 iv) 经典血浆蛋白质组，即约 400 种详细描述的蛋白质所有这些测定将提供明确的免疫功能的定量信息。为我们提供了一张精美的免疫状态图，这是一个假设，我们将通过关联我们的免疫状态来进行测试乙型肝炎疫苗接种前后测得的抗乙型肝炎滴度>500的结果样本将由波士顿儿童医院 NIAID 资助的 HIPC（人类）提供。免疫学项目联盟），这将显着增加该群体的影响力将样品备用的资助机会的要求提升到一个不同的水平，我们将推进我们的蛋白质组学流程，将吞吐量翻两番，达到每天至少 96 个样本并将血浆体积要求四分之一至 <100 纳升。因此，在未来，1微升血浆将足以进行数十次时间独立的LC/MS- 基于蛋白质测定，产生数百种蛋白质的定量信息，即使是来自最小和/或最虚弱患者的极少量血浆就足以建立任何人免疫状态的详细且公正的分子图谱。