Impact of tissue fixation on the isolation and detection of human brain proteome (DIA-LC-HRMS)

ABSTRACT: This study investigated the impact of tissue preservation methods on protein profiles analyzed by reversed-phase liquid chromatography-high-resolution mass spectrometry (LC-HRMS) using data-independent acquisition (DIA). We compared proteomic profiles from formalin-fixed, formalin-fixed and paraffin-embedded (FFPE), and fresh-frozen human brain tissues (cortex and hippocampus, n=6). Additionally, we evaluated an FFPE-specific commercial protein extraction kit (n=4). Formalin-fixed samples closely resembled fresh-frozen profiles, whereas FFPE samples exhibited greater deviations. A core set of 1,753 proteins was consistently detected across all sample preparation methods. A total of 35 proteins were identified exclusively in fresh-frozen samples but without functional enrichment. Quantitative comparisons to the proteome of fresh-frozen tissue revealed an underrepresentation of cellular processes, energy metabolism, signaling, and transport related to protein properties such as length, location, and hydrophobicity. In contrast, neuronal development and phagosome-related pathways were overrepresented in fixed tissues. In a pilot study comparing low (Braak 0-II, n=4) and high (Braak IV-VI, n=4) Alzheimer’s disease (AD) stages using formalin-fixed samples, we identified 12 potential protein biomarkers, primarily nucleosomal proteins and carboxypeptidase M (CPM). These findings suggest that formalin-fixed brain tissue provides reliable proteomic information, making it a valuable resource for neurodegenerative disease research.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Brain

DISEASE(S): Alzheimer's Disease

SUBMITTER: Laura Plantera  

LAB HEAD: Uta Ceglarek

PROVIDER: PXD064700 | Pride | 2026-03-16

REPOSITORIES: pride