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Ultrasensitive Top-Down Proteomics Analysis of Low Number of Mammalian Cells Using a Nanodroplet Sample Processing Platform

ABSTRACT: We developed an ultrasensitive top down platform by incorporating a microfluidic sample preparation system, termed nanoPOTS (Nanodroplet Processing in One pot for Trace Samples), into a top down workflow. We determined that a unique combination of a nonionic detergent DDM with urea as a protein extraction buffer can significantly improve both protein extraction efficiency and sample recovery. The nanoPOTS-based top down platform can reproducibly and quantitatively identify ~170 to ~620 proteoforms from ~70 to ~770 HeLa cells containing < ~100 ng total proteins. Data was searched with TopPIC and MSPathFinder.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Ying Zhu

PROVIDER: MSV000084612 | MassIVE | Fri Nov 22 10:53:00 GMT 2019

SECONDARY ACCESSION(S): PXD016410

REPOSITORIES: MassIVE

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Sensitive Top-Down Proteomics Analysis of a Low Number of Mammalian Cells Using a Nanodroplet Sample Processing Platform.

Zhou Mowei M Uwugiaren Naomi N Williams Sarah M SM Moore Ronald J RJ Zhao Rui R Goodlett David D Dapic Irena I Paša-Tolić Ljiljana L Zhu Ying Y

Analytical chemistry 20200506 10

Top-down proteomics is a powerful tool for characterizing genetic variations and post-translational modifications at intact protein level. However, one significant technical gap of top-down proteomics is the inability to analyze a low amount of biological samples, which limits its access to isolated rare cells, fine needle aspiration biopsies, and tissue substructures. Herein, we developed an ultrasensitive top-down platform by incorporating a microfluidic sample preparation system, termed nanoP ...[more]

PMID: 32374172

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