title: RNA-Seq of human platelets in Triple Negative PMF patients
design: Platelets Rich Plasma (PRP) was collected from fresh PB. PRP was diluted with an equal volume of platelet washing buffer supplemented with Apyrase (0.05U/ml), Hirudin (0.08U/ml) and Prostaglandin E1 (PGE1 0.3M). Diluted PRP was centrifuged at 450g for 5 to separate platelets (PLTs) from plasma. PLTs pellet was resuspended with 1.5ml PWB and filtered through a 5m filter to remove residual WBC. PLTs purification was based on the MACS Cell Separation technology to remove any residual WBC and RBC. PLTs were incubated with appropriate amounts of anti-CD235a and anti-CD45 antibodies and column-purified. PLTs were then lysed in 700l QIAzol Lysis Reagent and total RNA was extracted using the miRNeasy Micro Kit following manufacturers instructions.