A method for separating a volume of whole blood contained in a separation bag into at least three components comprising centrifuging the separation bag so as to separate therein a first inner layer comprising plasma, a second intermediate layer comprising platelets, a third intermediate layer comprising lymphocytes, monocytes and granulocytes and a fourth outer layer comprising red blood cells, transferring, into a plasma component bag connected to the separation bag, a plasma component and transferring into a mononuclear cell component bag a mononuclear cell components.