MassIVE MSV000091734

Complete Public PXD041621

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

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Description

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. [doi:10.25345/C5GT5FR46] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: LC-MS/MS based proteomics ; tandem mass tag ; limited sample preparation

Contact

Principal Investigators:
(in alphabetical order) 		Cheolju lee, Korea Institute of Science and Technology, south korea
Submitting User: nr_park

Publications

Park N, Lee H, Kwon Y, Ju S, Lee S, Yoo S, Park KS, Lee C.
One-STAGE Tip Method for TMT-Based Proteomic Analysis of a Minimal Amount of Cells.
ACS Omega. 2023 Jun 6;8(22):19741-19751. Epub 2023 May 24.

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