biostudies-literatureLee JYNational Institute of Environmental Health SciencesNIEHS NIH HHSUS Department of Energy Advanced Research Projects AgencyOffice of Biological and Environmental ResearchNational Institutes of HealthGenomic Science ProgramNational Institute of General Medical SciencesNIGMS NIH HHSPacific Northwest National Laboratory4193-4201https://www.ebi.ac.uk/biostudies/studies/S-EPMC9502155biostudies-literatureUnknown37(22)<h4>Motivation</h4>Ion mobility spectrometry (IMS) separations are increasingly used in conjunction with mass spectrometry (MS) for separation and characterization of ionized molecular species. Information obtained from IMS measurements includes the ion's collision cross section (CCS), which reflects its size and structure and constitutes a descriptor for distinguishing similar species in mixtures that cannot be separated using conventional approaches. Incorporating CCS into MS-based workflows can improve the specificity and confidence of molecular identification. At present, there is no automated, open-source pipeline for determining CCS of analyte ions in both targeted and untargeted fashion, and intensive user-assisted processing with vendor software and manual evaluation is often required.<h4>Results</h4>We present AutoCCS, an open-source software to rapidly determine CCS values from IMS-MS measurements. We conducted various IMS experiments in different formats to demonstrate the flexibility of AutoCCS for automated CCS calculation: (i) stepped-field methods for drift tube-based IMS (DTIMS), (ii) single-field methods for DTIMS (supporting two calibration methods: a standard and a new enhanced method) and (iii) linear calibration for Bruker timsTOF and non-linear calibration methods for traveling wave based-IMS in Waters Synapt and Structures for Lossless Ion Manipulations. We demonstrated that AutoCCS offers an accurate and reproducible determination of CCS for both standard and unknown analyte ions in various IMS-MS platforms, IMS-field methods, ionization modes and collision gases, without requiring manual processing.<h4>Availability and implementation</h4>https://github.com/PNNL-Comp-Mass-Spec/AutoCCS.<h4>Supplementary information</h4>Supplementary data are available at Bioinformatics online. Demo datasets are publicly available at MassIVE (Dataset ID: MSV000085979).Bioinformatics (Oxford, England)AutoCCS: automated collision cross-section calculation software for ion mobility spectrometry-mass spectrometry.PMC9502155P41 GM103493U2C ES030170U2CES03017014/CJ000/09/02DE-AC05-76RL01830P41 GM103493-15Zhou MZheng XBloodsworth KJIbrahim YMPayne SHSmith RDConant CRLee JYMetz TOFjeldsted JCOrton DJJansson CBilbao AHixson KKLi AWebb IKWilson JWfalseAutoCCS: automated collision cross-section calculation software for ion mobility spectrometry-mass spectrometry.<h4>Motivation</h4>Ion mobility spectrometry (IMS) separations are increasingly used in conjunction with mass spectrometry (MS) for separation and characterization of ionized molecular species. Information obtained from IMS measurements includes the ion's collision cross section (CCS), which reflects its size and structure and constitutes a descriptor for distinguishing similar species in mixtures that cannot be separated using conventional approaches. Incorporating CCS into MS-based workflows can improve the specificity and confidence of molecular identification. At present, there is no automated, open-source pipeline for determining CCS of analyte ions in both targeted and untargeted fashion, and intensive user-assisted processing with vendor software and manual evaluation is often required.<h4>Results</h4>We present AutoCCS, an open-source software to rapidly determine CCS values from IMS-MS measurements. We conducted various IMS experiments in different formats to demonstrate the flexibility of AutoCCS for automated CCS calculation: (i) stepped-field methods for drift tube-based IMS (DTIMS), (ii) single-field methods for DTIMS (supporting two calibration methods: a standard and a new enhanced method) and (iii) linear calibration for Bruker timsTOF and non-linear calibration methods for traveling wave based-IMS in Waters Synapt and Structures for Lossless Ion Manipulations. We demonstrated that AutoCCS offers an accurate and reproducible determination of CCS for both standard and unknown analyte ions in various IMS-MS platforms, IMS-field methods, ionization modes and collision gases, without requiring manual processing.<h4>Availability and implementation</h4>https://github.com/PNNL-Comp-Mass-Spec/AutoCCS.<h4>Supplementary information</h4>Supplementary data are available at Bioinformatics online. Demo datasets are publicly available at MassIVE (Dataset ID: MSV000085979).2021-01-01T00:00:00Z2021 Nov2025-04-19T03:34:05.935Z2025-04-07T13:42:31.973ZS-EPMC95021553414587410.1093/bioinformatics/btab429