biostudies-literatureUnknownRogers HTNHLBI NIH HHSTraining Program in Translational Cardiovascular ScienceNational Institutes of HealthNIGMS NIH HHSNIH HHSNational Science FoundationTop-down-mass spectrometry (MS)-based proteomics has emerged as a premier technology to examine proteins at the proteoform level, enabling characterization of genetic mutations, alternative splicing, and post-translational modifications. However, significant challenges that remain in top-down proteomics include the analysis of large proteoforms and the sensitivity required to examine proteoforms from minimal amounts of sample. To address these challenges, we have developed a new method termed "small-scale serial Size Exclusion Chromatography" (s3SEC), which incorporates a small-scale protein extraction (1 mg of tissue) and serial SEC without postfractionation sample handling, coupled with online high sensitivity capillary reversed-phase Analytical chemistryhttps://www.ebi.ac.uk/biostudies/studies/S-EPMC11298573biostudies-literatureSmall-Scale Serial Size Exclusion Chromatography (s3SEC) for High Sensitivity Top-Down Proteomics of Large Proteoforms.PMC11298573R01 HL109810T32 HL007936R01 GM125085DGE-1747503S10 OD018475R01 HL109810-06T32 HL007936-21R01 HL096971Rogers HTLarson EJFischer MSRossler KJGe YWang DGao ZAlpert AJMelby JAEhlers LEfalseSmall-Scale Serial Size Exclusion Chromatography (s3SEC) for High Sensitivity Top-Down Proteomics of Large Proteoforms.Top-down-mass spectrometry (MS)-based proteomics has emerged as a premier technology to examine proteins at the proteoform level, enabling characterization of genetic mutations, alternative splicing, and post-translational modifications. However, significant challenges that remain in top-down proteomics include the analysis of large proteoforms and the sensitivity required to examine proteoforms from minimal amounts of sample. To address these challenges, we have developed a new method termed "small-scale serial Size Exclusion Chromatography" (s3SEC), which incorporates a small-scale protein extraction (1 mg of tissue) and serial SEC without postfractionation sample handling, coupled with online high sensitivity capillary reversed-phase 2024-01-01T00:00:00Z2024 Feb2026-03-15T22:44:36.64Z2025-08-13T03:06:34.219ZS-EPMC112985733831563010.1021/acs.analchem.3c05733