In-Line Sample Processing System with an Immobilized Trypsin-Packed Fused-Silica Capillary Tube for the Proteomic Analysis of a Small Number of Mammalian Cells

Kosuke Hata, Yoshihiro Izumi, Takeshi Hara, Masaki Matsumoto, Takeshi Bamba

研究成果: ジャーナルへの寄稿記事

抜粋

Omics analysis at single-cell resolution has helped to demonstrate the shaping of cellular heterogeneity on the basis of the expression of various molecules. However, in-depth proteomic analysis of low-quantity samples has remained challenging because of difficulties associated with the measurement of large numbers of proteins by shotgun proteomics using nanoflow liquid chromatography tandem mass spectrometry (nano-LC/MS/MS). To meet such a demand, we developed a method called in-line sample preparation for efficient cellular proteomics (ISPEC) in which cells were captured, directly lysed, and digested with immobilized trypsin within fused-silica capillaries. ISPEC minimized sample loss during the sample preparation processes with a relatively small number of mammalian cells (<1000 cells) and improved the stability and efficiency of digestion by immobilized trypsin, compared to a conventional preparation method. Using our optimized ISPEC method with nano-LC/MS/MS analysis, we identified 1351, 351, and 60 proteins from 100 cells, 10 cells, and single cells, respectively. The linear response of the signal intensity of each peptide to the introduced cell number indicates the quantitative recovery of the proteome from a very small number of cells. Thus, our ISPEC strategy facilitates quantitative proteomic analysis of small cell populations.

元の言語英語
ページ(範囲)2997-3005
ページ数9
ジャーナルAnalytical chemistry
92
発行部数4
DOI
出版物ステータス出版済み - 2 18 2020

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All Science Journal Classification (ASJC) codes

  • Analytical Chemistry

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