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• W2965399948 abstract "Direct measurement of proteins produced by single cells promises to expand our understanding of molecular cell-to-cell differences (heterogeneity) and their contribution to normal and impaired development. High-resolution mass spectrometry (HRMS) is the modern technology of choice for the label-free identification and quantification of proteins, albeit usually in large populations of cells. Recent advances in microscale sample collection and processing, separation, and ionization have extended this powerful technology to single cells. This chapter describes a protocol based on microprobe capillary electrophoresis (CE) HRMS to enable the direct proteomic profiling of single cells embedded in complex tissues without the requirement for dissociation or whole-cell dissection. We here demonstrate the technology for identified individual cells in early developing embryos of Xenopus laevis and zebrafish as well as electrophysiologically identified single neurons in physiologically active brain slices from the mouse substantia nigra. Instructions are provided step-by-step to identify single cells using physiological or morphological cues, collect the content of the cells using microfabricated capillaries, and perform bottom-up proteomics using a custom-built CE electrospray ionization (ESI) mass spectrometer equipped with a quadrupole time-of-flight or orbitrap mass analyzer. Results obtained by this approach have revealed previously unknown differences between the proteomic state of embryonic cells and neurons. The data from single-cell proteomics by microprobe CE-ESI-HRMS complements those from single-cell transcriptomics, thereby opening exciting potentials to deepen our knowledge of molecular mechanisms governing cell and developmental processes." @default.
• W2965399948 created "2019-08-13" @default.
• W2965399948 creator A5001307632 @default.
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• W2965399948 date "2019-01-01" @default.
• W2965399948 modified "2023-10-16" @default.
• W2965399948 title "Single-cell proteomics in complex tissues using microprobe capillary electrophoresis mass spectrometry" @default.
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• W2965399948 doi "https://doi.org/10.1016/bs.mie.2019.07.001" @default.
• W2965399948 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/7397975" @default.
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