A Cost-Effective High-Throughput Plasma and Serum Proteomics Workflow Enables Mapping of the Molecular Impact of Total Pancreatectomy with Islet Autotransplantation,Journal of Proteome Research

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A Cost-Effective High-Throughput Plasma and Serum Proteomics Workflow Enables Mapping of the Molecular Impact of Total Pancreatectomy with Islet Autotransplantation
Journal of Proteome Research ( IF 3.8 ) Pub Date : 2018-04-19 , DOI: 10.1021/acs.jproteome.8b00111
Tue Bjerg Bennike _{1,

2,

3,

4} , Melena D. Bellin _{5,

6} , Yue Xuan ₇ , Allan Stensballe ₄ , Frederik Trier Møller ₈ , Gregory J. Beilman ₅ , Ofer Levy _{3,

9} , Zobeida Cruz-Monserrate ₁₀ , Vibeke Andersen _{11,

12} , Judith Steen ₁₃ , Darwin L. Conwell ₁₀ , Hanno Steen _{1,

2,

3}

Affiliation

Department of Pathology, Harvard Medical School, Boston, Massachusetts, United States
Department of Pathology, Boston Children’s Hospital, Boston, Massachusetts, United States
Precision Vaccines Program, Boston Children’s Hospital, Boston, Massachusetts, United States
Department of Health Science and Technology, Aalborg University, Aalborg, Denmark
Department of Surgery, University of Minnesota Medical Center, Minneapolis, Minnesota, United States
Department of Pediatrics, University of Minnesota Medical Center, Minneapolis, Minnesota, United States
Thermo Fisher Scientific, Bremen, Germany
Department of Epidemiology Research, Statens Serum Institut, Copenhagen, Denmark
Division of Infectious Diseases, Department of Medicine, Boston Children’s Hospital, Boston, Massachusetts, United States
Division of Gastroenterology, Hepatology and Nutrition, The Ohio State University Wexner Medical Center, Columbus, Ohio United States
Focused Research Unit for Molecular Diagnostic and Clinical Research (MOK), IRS-Center Sonderjylland, Hospital of Southern Jutland, Aabenraa, Denmark
Institute of Molecular Medicine, University of Southern Denmark, Odense, Denmark
F.M. Kirby Neurobiology Center, Boston Children’s Hospital, and Department of Neurology, Harvard Medical School, Boston, Massachusetts, United States

Blood is an ideal body fluid for the discovery or monitoring of diagnostic and prognostic protein biomarkers. However, discovering robust biomarkers requires the analysis of large numbers of samples to appropriately represent interindividual variability. To address this analytical challenge, we established a high-throughput and cost-effective proteomics workflow for accurate and comprehensive proteomics at an analytical depth applicable for clinical studies. For validation, we processed 1 μL each from 62 plasma samples in 96-well plates and analyzed the product by quantitative data-independent acquisition liquid chromatography/mass spectrometry; the data were queried using feature quantification with Spectronaut. To show the applicability of our workflow to serum, we analyzed a unique set of samples from 48 chronic pancreatitis patients, pre and post total pancreatectomy with islet autotransplantation (TPIAT) surgery. We identified 16 serum proteins with statistically significant abundance alterations, which represent a molecular signature distinct from that of chronic pancreatitis. In summary, we established a cost-efficient high-throughput workflow for comprehensive proteomics using PVDF-membrane-based digestion that is robust, automatable, and applicable to small plasma and serum volumes, e.g., finger stick. Application of this plasma/serum proteomics workflow resulted in the first mapping of the molecular implications of TPIAT on the serum proteome.

中文翻译：

具有成本效益的高通量血浆和血清蛋白质组学工作流程可实现胰岛自体移植对全胰腺切除术的分子影响的定位

血液是发现或监测诊断和预后蛋白质生物标记物的理想体液。然而，发现健壮的生物标志物需要分析大量样品以适当地代表个体间的变异性。为了解决这一分析难题，我们建立了高通量且具有成本效益的蛋白质组学工作流程，以在适用于临床研究的分析深度提供准确而全面的蛋白质组学。为了进行验证，我们从96孔板中的62个血浆样品中各提取了1μL，并通过独立于定量数据的采集液相色谱/质谱法分析了产物；使用Spectronaut进行特征量化来查询数据。为了显示我们的工作流程对血清的适用性，我们分析了来自48名慢性胰腺炎患者，胰岛自体移植（TPIAT）手术前后全胰腺切除术的一组独特样本。我们确定了16种具有统计学意义的丰度变化的血清蛋白，这些蛋白代表了不同于慢性胰腺炎的分子标记。总而言之，我们使用基于PVDF膜的消化建立了一种经济高效的高通量工作流程，用于全面的蛋白质组学研究，该消化过程坚固，可自动化执行，适用于少量血浆和血清，例如指棒。该血浆/血清蛋白质组学工作流程的应用导致了TPIAT对血清蛋白质组的分子影响的首次定位。

更新日期：2018-04-19

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