A Fully Integrated Spintip-Based Approach for Sensitive and Quantitative Profiling of Region-Resolved in Vivo Brain Glycoproteome

Peiwu Huang, Hongjie Li, Weina Gao, Zongwei CAI*, Ruijun Tian

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

7 Citations (Scopus)

Abstract

Region- and cell type-resolved global proteome and specific post-translational modifications (PTMs) profiling of tissues has drawn great attention recently for interpreting the heterogeneous multicellular microenvironment of various in vivo systems. Due to access to low microgram of proteins and low abundance of glycoproteins, spatially resolved glycoproteome analysis of in vivo tissue sections remains challenging. Several glycoproteomics sample preparation strategies were established for processing microgram-level of protein samples, but these strategies were not either fully integrated or directly compatible with tissue samples when considering protein extraction in strong lysis buffers. Moreover, these approaches mainly focused on identification of glycosylation sites, but pay less attention to quantification, all of which limit their applications. Here we designed a fully integrated spintip-based glycoproteomic approach (FISGlyco) which achieves all the steps of glycoprotein enrichment, digestion, deglycosylation, and desalting in single spintip device. Sample loss is significantly reduced, and the total processing time is reduced to 4 h, while detection sensitivity and label-free quantification precision is greatly improved. 607 N-glycosylation sites were successfully identified and quantified from only 5 μg of mouse brain proteins. By seamlessly combining with laser capture microdissection (LCM), the first region-resolved N-glycoproteome profiling of four mouse brain regions, including isocortex, hippocampus, thalamus, and hypothalamus, was achieved, with 1875, 1794, 1801, and 1417 N-glycosites identified, respectively. Our approach could be a generic approach for region and even cell type specific glycoproteome analysis of in vivo tissue sections.

Original languageEnglish
Pages (from-to)9181-9189
Number of pages9
JournalAnalytical Chemistry
Volume91
Issue number14
DOIs
Publication statusPublished - 17 Jun 2019

Scopus Subject Areas

  • Analytical Chemistry

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