Suspension TRAPping Filter (sTRAP) Sample Preparation for Quantitative Proteomics in the Low µg Input Range Using a Plasmid DNA Micro-Spin Column: Analysis of the Hippocampus from the 5xFAD Alzheimer’s Disease Mouse Model
Evangelia Thanou,
Frank Koopmans,
Débora Pita-Illobre,
Remco V. Klaassen,
Berna Özer,
Ioannis Charalampopoulos,
August B. Smit,
Ka Wan Li
Affiliations
Evangelia Thanou
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
Frank Koopmans
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
Débora Pita-Illobre
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
Remco V. Klaassen
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
Berna Özer
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
Ioannis Charalampopoulos
Pharmacology Department, Faculty of Medicine, University of Crete, 71003 Heraklion, Greece
August B. Smit
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
Ka Wan Li
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
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.
