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  English.dicp.cas.cn    Posted:2018-12-13
Lecture: In-Depth Proteome Profiling of Single Cells Including Circulating Tumor Cells using Nanodroplet Sample Processing and Ultrasensitive LC-MS

Time: 13th Dec., 2018, 9:30 am
Venue: Academic Hall, Biotechnology Building
Lecturer: Dr. Ryan T. Kelly, Brigham Young University

Introduction:

Dr. Ryan T. Kelly is Associate Professor of Brigham Young University, USA. He obtained his Ph.D. degree from Brigham Young University on 2005. From 2005 to 2007, he carried out Postdoctoral Research in Pacific Northwest National Laboratory, Richland, WA. From 2007 to 2010, he was appointed as Senior Research Scientist III, Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA. From 2010 to 2018, he is Senior Research Scientist IV, EMSL, Pacific Northwest National Laboratory, Richland, WA. From 2018 to now, he is Associate Professor of Department of Chemistry and Biochemistry, Brigham Young University, Provo, UT.

He is currently the Editorial Board Member of Scientific Reports, Nature Publishing Group. He was elected as councilor and chair of awards committee of American Electrophoresis Society from 2014 to 2017. Now he is the member of American Electrophoresis Society, CASSS Separations Society and American Chemical Society. He has published around more than 70 articles on famous scientific journals such as Angew. Chem. Int. Ed., Nat. Commun., Molecular & Cellular Proteomics, Chem. Sci. and Anal. Chem., written 15 books or book chapters and invented about 20 patens. He has gained many awards such as R&D 100 Award for “NanoPOTS: Nanodroplet Processing in One pot for Trace Samples” in 2018, R&D 100 Award for “Subambient Pressure Ionization with Nanoelectrospray (SPIN) Source” in 2015, Federal Laboratory Consortium Award for Excellence in Technology Transfer in 2012, Outstanding Performance Award, Fundamental and Computational Sciences Directorate, PNNL in 2010 and R&D 100 Award for “Ultrasensitive Electrospray Ionization Mass Spectrometry Source and Interface” in 2009.

Abstract:

Human tissues contain a variety of cell types and subtypes with distinct functions, and understanding heterogeneity at the single cell level is of great importance for biomedical research. Although MS-based proteomic analyses are capable of quantifying thousands of proteins, the extension to single cell studies has been largely ineffective. This is primarily due to protein and peptide losses during sample processing. To address this, we have developed nanoPOTS (Nanodroplet Processing in One-pot for Trace Samples) to efficiently process and analyze single mammalian cells. Cultured HeLa cells, primary lung cells, and spiked circulating tumor cells (CTCs) isolated from whole blood were FACS-sorted or isolated by laser microdissection intonanoPOTS chips based on cell morphology or immune his to chemistry staining. All processing steps were carried out in nanowells having a total processing volume <200 nL to reduce protein losses to surfaces. A custom robotic nanopipetting system was used to dispense reagents and collect prepared samples. The processed samples were analyzed by nanoLC/MS. We first used FACS-sorted HeLa cells to evaluate the sensitivity of present system. We are able to identify an average of 669 protein groups (n=3) per HeLa cell, with 332 quantifiable protein groups having valid LFQ values in at least 2 samples. We also tested whether the single cell proteomic platform could discriminate cell types isolated from human tissues. Antibody-labeled epithelial and fibroblast cells from the lung tissue of a healthy donor were used for the evaluation, and the two cell types were well clustered without overlap. Statistical analysis revealed a panel of proteins that were enriched for each cell type, which could serve as protein signatures. Finally, we analyzed spiked CTCs isolated from whole blood and could differentiate the single CTCs from white blood cells based on their proteomic signatures.

Contact: ZHANG LIhua, Group 1810
Phone: 84379720

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