Toward Analysis of Proteins in Single Cells: A Quantitative Approach Employing Isobaric Tags with MALDI Mass Spectrometry Realized with a Microfluidic Platform

Mian Yang, Randall Nelson, Alexandra Ros

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

Protein identification and quantification in individual cells is essential to understand biological processes such as those involved in cell apoptosis, cancer, biomarker discovery, disease diagnostics, pathology, or therapy. Compared with present single cell genome analysis, probing the protein content of single cells has been hampered by the lack of a protein amplification technique. Here, we report the development of a quantitative mass spectrometric approach combined with microfluidic technology reaching the detection sensitivity of high abundant proteins in single cells. A microfluidic platform with a series of chambers and valves, ensuring a set of defined wells for absolute quantification of targeted proteins, was developed and combined with isotopic labeling strategies employing isobaric tags for relative and absolute quantitation (iTRAQ)-labels. To this aim, we adapted iTRAQ labeling to an on-chip protocol. Simultaneous protein digestion and labeling performed on the microfluidic platform rendered the labeling strategy compatible with all necessary manipulation steps on-chip, including the matrix delivery for MALDI-TOF analysis. We demonstrate this approach with the apoptosis related protein Bcl-2 and quantitatively assess the number of Bcl-2 molecules detected. We anticipate that this approach will eventually allow quantification of protein expression on the single cell level.

Original languageEnglish (US)
Pages (from-to)6672-6679
Number of pages8
JournalAnalytical Chemistry
Volume88
Issue number13
DOIs
StatePublished - Jul 5 2016

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ASJC Scopus subject areas

  • Analytical Chemistry

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