Matrix-Assisted Ionization and Tandem Mass Spectrometry Capabilities in Protein Biomarker Characterization-An Initial Study Using the Small Cell Lung Cancer Biomarker Progastrin Releasing Peptide as a Model Compound.

Maren Christin Stillesby Levernæs,Cecilie Rosting,Trine Grønhaug Halvorsen

doi:10.1021/jasms.0c00336

Maren Christin Stillesby Levernæs, Cecilie Rosting + Show 1 more

Open Access

https://doi.org/10.1021/jasms.0c00336

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Abstract

This initial study evaluates vacuum matrix-assisted ionization (vMAI) mass spectrometry (MS) for identification and determination of tryptic peptides from the biomarker protein progastrin releasing peptide (ProGRP). Similar peptides and charge states were observed as in liquid chromatography (LC) electrospray ionization (ESI) MS. The prolonged ion duration in vMAI with similar charge states as in ESI was advantageous for determining the MS/MS fragmentation conditions compared to MAI. It is assumed that the vacuum ionization conditions lower the detection limits of the experiment. This may be the reason vMAI combined with high resolution MS enabled detection of tryptic peptides from more digested proteins than MAI selected reaction monitoring MS. Additionally, MAI ion mobility spectrometry MS (MAI-IMS-MS) was evaluated for differentiation of intact protein isoforms, successfully enabling differentiation of the isoforms by drift time selection. Examples are both shown for model proteins bovine serum albumin, cytochrome C, and lysozyme and the clinically relevant small cell lung cancer protein biomarker ProGRP, which exists in three isoforms. Coupling with the vacuum ionization conditions using a dedicated vacuum-probe source MAI enables information to be extracted readily as with conventional approaches, just faster.

Highlights

In recent years, mass spectrometry (MS) has been gaining acceptance in clinical environments
Due to limited availability of the progastrin releasing peptide (ProGRP) isoforms, initial method development was performed with well-known standard proteins (bovine serum albumin (BSA), lysozyme, and cytochrome C) and their tryptic digests. vacuum matrix-assisted ionization (vMAI)-MS analyses of the tryptic digests of BSA, lysozyme, and cytochrome C showed long ion abundances (4−5 min) but moderate sequence coverage (39−54%)
For BSA, the coverage was substantially lower than when analyzing a BSA digest by liquid chromatography (LC)− electrospray ionization (ESI)-MS, probably due to the low ion abundances

Summary

Introduction

Mass spectrometry (MS) has been gaining acceptance in clinical environments. The simplicity of matrix-assisted ionization (MAI) MS requiring neither a high voltage source, heat, laser, nor ion beam makes it an attractive technique for development of low cost, user-friendly, and high throughput solutions for the clinical laboratory.[5] we have recently implemented MAI-MS in our lab to, cautiously, explore its use in targeted bottom-up analysis of proteins.[6] In this work, previously established liquid chromatography electrospray ionization selected reaction monitoring (LC-ESI-SRM)−MS transitions of tryptic signature peptides were applied in targeted MAISRM-MS analysis from biological matrices.[6]

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