Molecular profiling of vascular beds in vivo by DISDIVO
We developed a new technique named as Differential Systemic Decellularization in vivo (DISDIVO) for the molecular profiling of vascular beds throughout the body. This method can by applied to murine models. Thus, the data provided here correspond to mice vascular beds obtained in vivo by DISDIVO.
Sample Processing Protocol
Biological samples obtaned from DISDIVO were processed for LC-MS/MS by reducing the disulfide bonds and doing an alkylation with iodoacetamide. After alkylation proteins were digested using trypsin. Samples were subsequently desalted using a C18 cartridge and each sample was fractionated by high-pressure liquid chromatography using a C18 column. Samples were then combined and dried in a vacuum concentrator. Finally, samples were resuspended and analyzed by LC-MS/MS using a Dionex Ultimate 3000 UHPLC system coupled with an Orbitrap Elite mass spectrometer. Fragmentation of ions was performed using high energy collision dissociation fragmentation mode.
Data Processing Protocol
Database searching of raw proteomics data was performed using PEAKS Studio v7.5 with 10 ppm of precursor ion tolerance and 0.05 Da of fragment ion tolerance. False discovery rate was set at 1%. Carbamidomethylation (C) was set as fixed modification and other post-translational modifications were searched automatically using PEAKS PTM algorithm. The uniprot mouse database (58761 entries, downloaded on 18/02/2016) was used for searching.
Aida Serra, School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551.
Siu Kwan Sze, School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551, Singapore ( lab head )