Isolation of whole mononuclear cells from peripheral blood and cord blood

Curr Protoc Immunol. 2009 Apr;Chapter 7:Unit7.1. doi: 10.1002/0471142735.im0701s85.


Peripheral blood is the primary source of lymphoid cells for investigation of the human immune system. Its use is facilitated by Ficoll-Hypaque density gradient centrifugation-a simple and rapid method of purifying peripheral blood mononuclear cells (PBMC) that takes advantage of the density differences between mononuclear cells and other elements found in the blood sample. Thus, cells are distributed in the solution in layers based on the differences in their density/size. Additional purification methods can be employed as the mononuclear cell sample can be purified from monocytes by adherence or by exposure to L-leucine methyl ester; these methods are described for both procedures. Cord blood and peripheral blood from infants contain immature cells, including nucleated red cells, which can result in significant contamination of the mononuclear cell layer, and removal of these cells requires additional steps that are described. The isolation procedures presented here can also be applied to cell populations derived from tissues.

MeSH terms

  • Cell Separation* / instrumentation
  • Cell Separation* / methods
  • Centrifugation, Density Gradient
  • Erythroblasts / cytology
  • Fetal Blood / cytology*
  • Ficoll
  • Humans
  • Immunologic Techniques
  • Infant, Newborn
  • Leucine
  • Leukocytes, Mononuclear / cytology*
  • Macrophages / cytology
  • Methyl Ethers


  • Methyl Ethers
  • Ficoll
  • Leucine