Dxh 9000 hematology analyzer

PBMCs isolation was carried out using Ficoll-Paque following the manufacturer instructions with some minor modifications. Briefly, blood samples were collected by venipuncture into 8 mL ACD tubes and centrifuged at 300g for 10 min to allow plasma collection. A volume of PBS, equal to the volume of plasma collected, was then added to each tube and resulting blood samples were placed onto a layer of Ficoll-Paque (blood/Ficoll-Paque ratio of 4:3). Afterwards, samples were centrifuged at 500g for 30 min. PBMCs were subsequently collected, washed two times with PBS and counted on a flow cytometer (DXH-9000 hematology analyzer, Beckman Coulter^®).

Blood samples were collected by venipuncture into ACD tubes (acid citric dextrose, BD Vacutainer^®). All subsequent steps were performed at room temperature. PBMCs were subsequently extracted by density gradient using Ficoll-Paque (Ge Healthcare^®) following the manufacturer protocol with minor modifications. Briefly, blood samples were centrifuged at 300g for 10 minutes to allow separation and collection of plasma rich platelets (PRP). A volume of PBS (phosphate buffer saline), equal to the volume of plasma collected, was then added to each tube. The blood samples were carefully place onto a layer of Ficoll-Paque (blood:Ficoll-Paque ratio of 4:3) and centrifuged at 500g for 30 minutes. PBMCs were thereafter isolated form the interface and collected by centrifugation (500 g, 10 minutes). Cell pellets were wash twice PBS and counted on a DXH-9000 hematology analyzer (Beckman Coulter^®).