PBMC isolation from whole blood

Leucocytes/PBMCs isolation from whole blood



  • Separating solution should be at RT
  • Pipette 15ml separating solution into Leucosep-tube
  • Centrifuge tube with separating solution 30sec 1000g at RT
  • Dilute 10ml blood 1:2 with 0.9 % sodium chloride solution, final volume 30ml


  • Add carefully blood solution to the Leucosep tube (separating solution is already inside)
  • Centrifuge 15min 800g at RT without brake!
  • Pipette intermediate phase (leucocytes/PBMCs) into a new 15ml tube (lower phase (cell pellet): erythrocytes/granulocytes, middle phase: leucocytes/PBMCs (above the filter and separating solution), upper phase: plasma
  • Wash cells with 10ml PBS 10min 250g at RT, remove supernatant
  • Resuspend cell pellet with 5ml PBS, centrifuge 10min 250g at RT, remove supernatant
  • Repeat step 5


Leucosep-tube, sterile, 50ml Greiner Bio-One #227290
Biocoll separating solution Biochrom AG #L6115
0.9 % sodium chloride solution