Dataset Information

One-STAGE Tip Method for TMT based Proteomic Analysis of Minimal Amount of Cells

ABSTRACT: LC-MS-based profiling of proteomes with isobaric tag labeling from low-quantity biological and clinical samples, including needle-core biopsies and laser capture microdissection, has been challenging due to limited amount and sample loss during preparation. To address this problem, we developed OnM (On-Column from Myers et al. and mPOP) modified On-column method combining freeze-thaw lysis of mPOP with isobaric tag labeling of On-Column method to minimize sample loss. OnM is a method that processes the sample in one STAGE-tip from cell lysis to Tandem Mass Tag (TMT) labeling without any transferring of the sample. In terms of protein coverage, cellular components, and TMT labeling efficiency, modified On-column (or OnM) displayed similar performance to the results from Myers et al. To evaluate the lower-limit processing capability of OnM, we utilized OnM for multiplexing and were able to quantify 301 proteins in a TMT 9-plex with 50 cells per channel. We optimized the method as low as 5 cells per channel in which we identified 51 quantifiable proteins. OnM method is a low-input proteomics method widely applicable and capable of identifying and quantifying proteomes from limited samples with tools that are readily available in majority of proteomic laboratories.

INSTRUMENT(S): Q Exactive

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

SUBMITTER: Cheolju lee

PROVIDER: MSV000091734 | MassIVE | Tue Apr 18 04:04:00 BST 2023

SECONDARY ACCESSION(S): PXD041621

REPOSITORIES: MassIVE

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One-STAGE Tip Method for TMT-Based Proteomic Analysis of a Minimal Amount of Cells.

Park Narae N Lee Hankyul H Kwon Yumi Y Ju Shinyeong S Lee Seonjeong S Yoo Seongjin S Park Kang-Sik KS Lee Cheolju C

ACS omega 20230524 22

Liquid chromatography-tandem mass spectrometry (LC-MS)-based profiling of proteomes with isobaric tag labeling from low-quantity biological and clinical samples, including needle-core biopsies and laser capture microdissection, has been challenging due to the limited amount and sample loss during preparation. To address this problem, we developed OnM (On-Column from Myers et al. and mPOP)-modified on-column method combining freeze-thaw lysis of mPOP with isobaric tag labeling of On-Column method ...[more]

PMID: 37305273

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