Tapered-Tip Capillary Electrophoresis Nano-Electrospray Ionization Mass Spectrometry for Ultrasensitive Proteomics: the Mouse Cortex

  • Sam B. Choi
  • Marta Zamarbide
  • M. Chiara Manzini
  • Peter Nemes
Focus: Emerging Investigators: Research Article
  • 436 Downloads

Abstract

Ultrasensitive characterization of the proteome raises the potential to understand how differential gene expression orchestrates cell heterogeneity in the brain. Here, we report a microanalytical capillary electrophoresis nano-flow electrospray ionization (CE-nanoESI) interface for mass spectrometry to enable the measurement of limited amounts of proteins in the mouse cortex. Our design integrates a custom-built CE system to a tapered-tip metal emitter in a co-axial sheath-flow configuration. This interface can be constructed in <15 min using readily available components, facilitating broad adaptation. Tapered-tip CE-nanoESI generates stable electrospray by reproducibly anchoring the Taylor cone, minimizes sample dilution in the ion source, and ensures efficient ion generation by sustaining the cone-jet spraying regime. Parallel reaction monitoring provided a 260-zmol lower limit of detection for angiotensin II (156,000 copies). CE was able to resolve a complex mixture of peptides in ~330,000 theoretical plates and identify ~15 amol (~1 pg) of BSA or cytochrome c. Over 30 min of separation, 1 ng protein digest from the mouse cortex yielded 217 nonredundant proteins encompassing a ~3-log-order concentration range using a quadrupole time-of-flight mass spectrometer. Identified proteins included many products from genes that are traditionally used to mark oligodendrocytes, astrocytes, and microglia. Finally, key proteins involved in neurodegenerative disorders were detected (e.g., parkinsonism and spastic paraplegia). CE-nanoESI-HRMS delivers sufficient sensitivity to detect proteins in limited amounts of tissues and cell populations to help understand how gene expression differences maintain cell heterogeneity in the brain.

Graphical Abstract

Keywords

Capillary electrophoresis Mass spectrometry Bottom-up proteomics Neuron Central nervous system Mouse 

Supplementary material

13361_2016_1532_MOESM1_ESM.xlsx (133 kb)
ESM 1(XLSX 132 kb)
13361_2016_1532_MOESM2_ESM.xlsx (215 kb)
ESM 2(XLSX 215 kb)

Copyright information

© American Society for Mass Spectrometry 2016

Authors and Affiliations

  1. 1.Department of ChemistryThe George Washington UniversityWashingtonUSA
  2. 2.Department of Pharmacology and PhysiologyThe George Washington UniversityWashingtonUSA

Personalised recommendations