iProXProteomicsYun YangHomo SapiensMus Musculushttp://www.iprox.org/page/project.html?id=IPX0013448000yangyunwww@126.comInternational Academy of Phronesis Medicine (Guang Dong)iProXLarge-scale tissue proteomics requires workflows that are efficient, rapid, and repeatable across diverse samples. Herein, we present a Simple Workflow for Integrated and Fast Tissue-preparation (SWIFT), which enables complete processing of fresh-frozen (FF) and formalin-fixed, paraffin-embedded (FFPE) tissues in either one- or two-step formats while maintaining deep proteome coverage with high repeatability from low- to microgram-level tissues. For FF tissues, an incubation process integrating lysis, reduction, alkylation, and digestion generates peptide samples directly from tissues in ≤1.5 h. For FFPE tissues, concurrent deparaffinization, rehydration, and de-cross-linking are achieved within 0.5 h, followed by one-step peptide preparation. Furthermore, our workflows eliminate desalting and offline cleanup steps, thereby reducing variability and total processing time. Using our methods, we identified up to ∼10,000 protein groups and ∼150,000 peptides across multiple mouse organs on the timsTOF Pro. Repeatability was high (pairwise Pearson's <i>r</i> > 0.96 across six experimental replicates), with dynamic ranges spanning 6-7 orders of magnitude. Organ-enriched protein analysis identified functionally distinct proteins unique to each tissue. Paired FF and FFPE analyses revealed preservation-induced shifts, with FFPE tissues showing reduced detection of membrane-associated and respiratory proteins, including mitochondrial Complex I. Together, our fast and simplified workflows enable deep tissue proteomics for large-scale clinical and translational studies in a cost-effective and widely accessible manner.Simple, Fast, and Highly Efficient One- or Two-Step Proteomic Preparation Enables Deep Profiling of Microgram-Level FF and FFPE Tissues.Wei Chuping C, Zhang Qiuxia Q, Fu Changying C, Leng Yeye Y, Huang Chuanxi C, He Fuchu F, Yang Yun YfalseSimple Workflow for Integrated and Fast Tissue-preparation for microgram-level FF and FFPE Tissues Proteomics AnalysisWe present Simple Workflow for Integrated and Fast Tissue-preparation (SWIFT) workflows, enabling complete processing of fresh-frozen (FF) and formalin-fixed, paraffin-embedded (FFPE) tissues in one- or two-step formats, while maintaining deep proteome coverage with good repeatability. For FF tissues, an incubation process integrating lysis, reduction, alkylation, and digestion can generate peptide samples directly from tissues in ≤ 1.5 h. For FFPE tissues, simultaneous deparaffinization, rehydration, and de-crosslinking was achieved within 0.5 h, followed by one-step pretreatment to obtain peptides.Mon Apr 27 00:00:00 BST 2026PXD07767910090960642046375