Dataset Information

A Tiered Approach to Human Synapse Proteomics: Optimized LC-MS/MS Analysis of Whole-Tissue Lysate and Synaptosome Preparations from Frozen Post-Mortem Brain Samples

ABSTRACT: Recent advancements in neuroproteomics have enabled detailed analysis of protein expression and function in the human brain, improving our understanding of the relationship between genetics, cell biology of neurological and psychiatric disorders and their clinical diagnosis. Synaptic dysfunction often has a central role in these disorders. Leveraging the high sensitivity of modern liquid chromatography-tandem mass spectrometry (LC-MS/MS), we here evaluated the detection of synaptic proteins in whole-tissue lysates compared to enriched synaptosome preparations. First, we optimized sample preparation protocols for frozen human gray matter (GM), refining the suspension TRAPping (sTRAP) digestion method to improve protein solubilization and Cysteine reduction and alkylation using thin human tissue sections, and to accomplish low technical variation by minimizing sample handling. We achieved a highly reproducible sample preparation workflow by rigorously applying standardization and randomization across dissection, processing, and LC-MS/MS runs. Second, comparative LC-MS/MS analysis showed that cortical whole-tissue lysates are a practical solution for large-scale studies and broadly detected synaptic proteins. However, synaptosome isolation offered improved resolution of synapse-specific proteins. Because synapse-proteomics enables insight into spatial regulation—i.e., alterations at the synapse not reflected in the soma– we recommend a tiered approach for large-scale studies: initial whole-tissue lysate analysis for broad disease-associated changes, followed by targeted synaptosome proteomics to deepen insight into synaptic alterations. This strategy optimally balances throughput, reproducibility, and biological relevance, and enhances the study of brain disorders through proteomics. Moreover, analyzing synaptic proteins first at the tissue level improves insight into overall regulation of synaptic proteins induced by synapse loss or gain.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Brain

SUBMITTER: Remco Klaassen

LAB HEAD: Sabine Spijker

PROVIDER: PXD076278 | Pride | 2026-05-25

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	20231109_fkuhn_Tissue-extraction-pilot_DIA_55C_NBR_Post_A-1_S1-A5_1_11377.d.zip	Other
	20231109_fkuhn_Tissue-extraction-pilot_DIA_55C_NBR_Post_A-2_S1-A6_1_11409.d.zip	Other
	20231109_fkuhn_Tissue-extraction-pilot_DIA_55C_NBR_Post_A-3_S1-A7_1_11361.d.zip	Other
	20231109_fkuhn_Tissue-extraction-pilot_DIA_55C_NBR_Post_A-4_S1-A8_1_11393.d.zip	Other
	20231109_fkuhn_Tissue-extraction-pilot_DIA_55C_NBR_Post_B-1_S1-C5_1_11385.d.zip	Other

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A Tiered Approach to Human Synapse Proteomics: Optimized LC-MS/MS Analysis of Whole-Tissue Lysate and Synaptosome Preparations from Frozen Post-Mortem Brain Samples.

Roig-Kuhn Femke C FC Klaassen Remco V RV Koopmans Frank T W FTW Koolman Tiara S Z TSZ Smit August B AB Spijker Sabine S

Cells 20260421 8

Recent advancements in neuroproteomics have enabled detailed analysis of protein expression in the human brain, yet resolving synaptic dysfunction-a central feature of many neurological and psychiatric disorders-requires careful methodological consideration. Leveraging the high sensitivity of modern liquid chromatography-tandem mass spectrometry (LC-MS/MS), we evaluated the utility of whole-tissue lysates versus enriched synaptosome preparations for detecting synaptic protein signatures. First, ...[more]

PMID: 42041603

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