# Chromatographic and Electrophoretic Separations Optimized for Native MS

> **NIH NIH P41** · OHIO STATE UNIVERSITY · 2020 · $129,026

## Abstract

TR&D 4 Project Summary. Innovative and cutting-edge separation and ionization strategies are integral to
continue advancing mass spectrometry (MS) as an analytical tool in biomedical research. This is evidenced by
efforts to make proteolytically digested samples a more accessible application for protein identification and
quantification (bottom-up approach) and through the continued development of one- and multi-dimensional
separation methods for intact proteins, which ultimately lead to in-depth sequence determination of thousands
of proteins and protein isoforms within a single sample (top-down approach). Most native MS (nMS) applications
center on the characterization of a limited set of proteins and protein complexes in samples of low complexity.
This constraint is attributed to the use of direct sample infusion via nano electrospray ionization (nanoESI) for
nMS of 'pure' protein preparations, which is limited by the need to generate a sufficient quality and quantity of
the targeted protein prior to nMS. Ultimately these efforts are labor, time, and cost intensive. The overarching
goal of TR&D 4 is to develop cutting-edge separation technologies in order to significantly advance
research in native proteins fundamental to human health, and driven by biological problems. The
innovation, maturation, and dissemination of technologies with a higher throughput and lower cost than currently
possible will lead to a stronger understanding of a larger set of protein complexes. Specifically, we will develop
i) one- and two-dimensional column chromatographic methods with the additional possibility for charge
manipulation during electrospray ionization, ii) one- and two-dimensional fiber plate separation methods in
conjunction with desorption electrospray ionization and paper spray-like ionization, and iii) tunable capillary
electrophoresis methods with unprecedented selectivity, compatible with native mass spectrometry. A strong
cadre of ionization/ separation experts with well-developed ionization techniques (PI: Badu), novel
chromatographic stationary phases (PI: Olesik), enhanced fluids (PI: Olesik), smart materials with tunable
capillary electrophoresis separations (PI: Holland), and nMS (PI: Wysocki) is represented. These efforts leverage
collaborations with Phenomenex and Sciex, two leading separation industry partners. Access to multiple soluble
protein complexes (DBP 1-3A, DBP 6-10) and membrane proteins (DBP 3B-5) serve as an authentic test-bed to
drive separation methods and ionization techniques that address critical barriers faced by prominent biomedical
researchers in the field. The TR&D 4 separation and ionization methodologies are synergistic with improvements
in surface-induced dissociation (TR&D 1), ion mobility measurements (TR&D 2), and hybrid methods for
complex-down analysis (TR&D 3), accessible for a greater number of samples from all areas of protein related
research. In combination with software developments to automate data analy...

## Key facts

- **NIH application ID:** 9978849
- **Project number:** 5P41GM128577-03
- **Recipient organization:** OHIO STATE UNIVERSITY
- **Principal Investigator:** Susan V. Olesik
- **Activity code:** P41 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $129,026
- **Award type:** 5
- **Project period:** — → —

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9978849

## Citation

> US National Institutes of Health, RePORTER application 9978849, Chromatographic and Electrophoretic Separations Optimized for Native MS (5P41GM128577-03). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9978849. Licensed CC0.

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