Updated publication reference for PubMed record(s): 29323256. Platelets are major players in the process of intravascular thrombus formation. Current therapeutic strategies still fail to prevent thrombotic coronary events in a substantial number of patients, indicating that the complex mechanisms modulating platelet response during activation are not fully elucidated. The evidence that platelets are capable of de novo protein synthesis has raised the issue of whether and how these resident mRNAs are regulated in circulating platelets. Among the several mechanisms potentially involved, mRNA splicing may be relevant. Purified platelet-rich plasmas from healthy volunteers were collected and in vitro activated with collagen or Thrombin Receptor Activating Peptide (TRAP). Transcriptome profiling by RNA-Seq and in silico intron representation analysis were applied to search for the presence of pre-mRNA molecules and splicing events affected by platelet activation. HiRIEF LC-MS allowed platelet proteome characterization at deep coverage to investigate a possible correlation between splicing events and protein levels. By comparing computational and wet-lab analyses it was possible to identify a set of transcripts influenced at both intron and protein level.