biostudies-literatureScheid JCHIST-ERADeutsche ForschungsgemeinschaftFonds Wetenschappelijk Onderzoek,BelgiumHORIZON EUROPE Project BAXERNA 2.0Invest BW Innovation grantDeutsche KrebshilfeElse Kröner-Fresenius-Stiftung290https://www.ebi.ac.uk/biostudies/studies/S-EPMC12455830biostudies-literatureUnknown26(1)Confident identification of human leukocyte antigen (HLA)-presented peptides is crucial for advancing cancer immunotherapy. We present MHCquant2, a scalable and modular Nextflow pipeline integrated into nf-core as a reproducible, portable, and standardized workflow for immunopeptidomics. This integration allows a community-driven, robust solution for high-throughput analyses across operating systems and cloud infrastructures. MHCquant2 integrates open-source tools including OpenMS, DeepLC, and MS²PIP, improving peptide identifications by up to 27% across diverse MS platforms, particularly enriching low-abundant peptides. MHCquant2 demonstrates state-of-the-art performance on our novel benign_MHCquant2 dataset (n = 92) and expands the benign human immunopeptidome by oveGenome biologyMHCquant2 refines immunopeptidomics tumor antigen discovery.PMC124558302022_EKSE.79EXC2180-390900677BW1_4064/03/TruVac70114948ODEEP-EU10108054412B7123N, G010023N, G028821N, 1SE3724NMartens LLemke SSachsenberg TDeclerq AHoenisch-Gravel NWacker MGabriels RBichmann LDubbelaar MLBauer JScheid JDengler ANahnsen SWalz JSfalseMHCquant2 refines immunopeptidomics tumor antigen discovery.Confident identification of human leukocyte antigen (HLA)-presented peptides is crucial for advancing cancer immunotherapy. We present MHCquant2, a scalable and modular Nextflow pipeline integrated into nf-core as a reproducible, portable, and standardized workflow for immunopeptidomics. This integration allows a community-driven, robust solution for high-throughput analyses across operating systems and cloud infrastructures. MHCquant2 integrates open-source tools including OpenMS, DeepLC, and MS²PIP, improving peptide identifications by up to 27% across diverse MS platforms, particularly enriching low-abundant peptides. MHCquant2 demonstrates state-of-the-art performance on our novel benign_MHCquant2 dataset (n = 92) and expands the benign human immunopeptidome by ove2025-01-01T00:00:00Z2025 Sep2026-06-03T19:40:25.585Z2026-05-01T03:10:19.253ZS-EPMC124558304098392510.1186/s13059-025-03763-8