Department of Molecular and Cellular Neurobiology, Center for Neurogenomics and Cognitive Research, Amsterdam Neuroscience, Vrije Universiteit Amsterdam, De Boelelaan 1085, 1081 HV Amsterdam, The Netherlands.
Pharmacology Department, Faculty of Medicine, University of Crete, 71003 Heraklion, Greece.
Cells. 2023 Apr 25;12(9):1242. doi: 10.3390/cells12091242.
Suspension TRAPping filter (sTRAP) is an attractive sample preparation method for proteomics studies. The sTRAP protocol uses 5% SDS that maximizes protein solubilization. Proteins are trapped on a borosilicate glass membrane filter, where SDS is subsequently removed from the filter. After trypsin digestion, peptides are analyzed directly by LC-MS. Here, we demonstrated the use of a low-cost plasmid DNA micro-spin column for the sTRAP sample preparation of a dilution series of a synapse-enriched sample with a range of 10-0.3 µg. With 120 ng tryptic peptides loaded onto the Evosep LC system coupled to timsTOF Pro 2 mass spectrometer, we identified 5700 protein groups with 4% coefficient of variation (CoV). Comparing other sample preparation protocols, such as the in-gel digestion and the commercial Protifi S-TRAP with the plasmid DNA micro-spin column, the last is superior in both protein and peptide identification numbers and CoV. We applied sTRAP for the analysis of the hippocampal proteome from the 5xFAD mouse model of Alzheimer's disease and their wildtype littermates, and revealed 121 up- and 54 down-regulated proteins. Protein changes in the mutant mice point to the alteration of processes related to the immune system and Amyloid aggregation, which correlates well with the known major Alzheimer's-disease-related pathology. Data are available via ProteomeXchange with the identifier PXD041045.
悬浮陷阱过滤（sTRAP）是一种有吸引力的蛋白质组学研究样品制备方法。sTRAP 方案使用 5% SDS，可最大限度地提高蛋白质的溶解。蛋白质被捕获在硼硅酸盐玻璃膜过滤器上，随后 SDS 从过滤器中去除。经过胰蛋白酶消化后，肽直接通过 LC-MS 进行分析。在这里，我们展示了使用低成本质粒 DNA 微柱用于 sTRAP 样品制备一系列突触富集样品的稀释系列，范围为 10-0.3 µg。将 120 ng 胰蛋白酶肽加载到与 timsTOF Pro 2 质谱仪相连的 Evosep LC 系统上，我们鉴定了 5700 个蛋白组，变异系数（CoV）为 4%。与其他样品制备方案（如胶内消化和商业 Protifi S-TRAP 与质粒 DNA 微柱）相比，最后一种方案在蛋白和肽鉴定数量和 CoV 方面均具有优势。我们应用 sTRAP 分析了阿尔茨海默病 5xFAD 小鼠模型及其野生型同窝仔鼠的海马蛋白质组，发现了 121 个上调和 54 个下调蛋白。突变小鼠中的蛋白变化表明与免疫系统和淀粉样蛋白聚集相关的过程发生了改变，这与已知的主要阿尔茨海默病相关病理学密切相关。数据可通过 ProteomeXchange 以标识符 PXD041045 获得。
