Evaluating the Effects of Acquisition and Search Parameters on Repository-Scale MS/MS Matching
- 1Department of Chemistry, Virginia Tech, Blacksburg, Virginia 24061, United States
Abstract
Tandem mass spectrometry (MS/MS) spectral matching is increasingly performed at the repository scale, where tools such as MASST enable a query spectrum to be compared against heterogeneous public data. Unlike analogous genomic-based searches, repository-scale MS/MS matching lacks integrated false discovery rate (FDR) reporting, leaving parameter selection entirely to user discretion. Here, we evaluate how acquisition factors and analysis filters jointly affect MS/MS matching using a curated library of 40 metabolites spanning ten chemical classes. In this study, we find that normalized collision energies from 30 to 50% balance sufficient fragmentation against loss of larger diagnostic ions, producing the most structurally informative spectra and yielding the highest proportion of repository matches. Using a fragmentation-based decoy library, we show that six minimum matching peaks achieve <1% FDR across the entire cosine threshold range tested (0.7–1.0), while fewer matching peaks require increasingly stringent cosine thresholds. These findings provide practical, benchmarking guidance for user-provided acquisition and search parameters in MASST searches.
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BibTeX
@article{keys2026evaluating,
title = {{Evaluating the Effects of Acquisition and Search Parameters on Repository-Scale MS/MS Matching}},
author = {Allison J. Keys and Emily C. Gentry},
journal = {Journal of the American Society for Mass Spectrometry},
year = {2026},
month = oct,
doi = {10.1021/jasms.6c00332},
url = {https://doi.org/10.1021/jasms.6c00332},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Evaluating the Effects of Acquisition and Search Parameters on Repository-Scale MS/MS Matching
AU - Allison J. Keys
AU - Emily C. Gentry
PY - 2026
DA - 2026/10/23
JO - Journal of the American Society for Mass Spectrometry
DO - 10.1021/jasms.6c00332
UR - https://doi.org/10.1021/jasms.6c00332
AB - Tandem mass spectrometry (MS/MS) spectral matching is increasingly performed at the repository scale, where tools such as MASST enable a query spectrum to be compared against heterogeneous public data. Unlike analogous genomic-based searches, repository-scale MS/MS matching lacks integrated false discovery rate (FDR) reporting, leaving parameter selection entirely to user discretion. Here, we evaluate how acquisition factors and analysis filters jointly affect MS/MS matching using a curated library of 40 metabolites spanning ten chemical classes. In this study, we find that normalized collision energies from 30 to 50% balance sufficient fragmentation against loss of larger diagnostic ions, producing the most structurally informative spectra and yielding the highest proportion of repository matches. Using a fragmentation-based decoy library, we show that six minimum matching peaks achieve <1% FDR across the entire cosine threshold range tested (0.7–1.0), while fewer matching peaks require increasingly stringent cosine thresholds. These findings provide practical, benchmarking guidance for user-provided acquisition and search parameters in MASST searches.
SN - 1044-0305
SN - 1879-1123
ER -