Density Gradient Media – News and Views

Flotation of monocytes from a leukocyte-rich fraction through a simple discontinuous iodixanol gradient provides a simple method for their purification. Both Graziani-Bowering et al [1] and later Nutt et al [2] found that the purity based on CD14/CD4 [1] or CD68 [2] analysis was at least 90%. Nutt et al [2] compared the monocyte isolates with those from centrifugal elutriation and concluded that they were comparable. The method is easy, rapid and avoids the use of special expensive rotor hardware. More recently [3] it has been recommended in studies on monocyte-derived macrophages.

Yields by this flotation method, which is carried out at room temperature, are approx. 35%. Recently an alternative even more rapid flotation method was developed. It uses whole blood and is carried out at a lower temperature; the yields are raised to approx 60%, but the purity is reduced to approx. 80%.

Detailed protocols of both monocyte isolation strategies, from a leukocyte-rich plasma (Application Sheet C09) and from whole blood (Application Sheet C10) and abstracts of papers reporting the use of these methods may be accessed from the Cell Index on the Axis-Shield website; click HERE and follow the instructions.

1 Graziani-Bowering, G.M., Graham, J. and Filion, L.G. (1997) A quick, easy and inexpensive method for the isolation of human peripheral blood monocytes J. Immunol. Methods, 207, 157-168

2 Nutt, J.C., Willis, C.C., Morris, J.M. and Gallery, E.D.M. (2004) Isolating pure populations of monocytes from the blood of pregnant women: comparison of flotation in iodixanol with elutriation J. Immunol. Methods, 293, 215-218

3 Quesniaux, V., Erard, F. and Ryffel, B. (2010) Adjuvant activity on murine and human macrophages In Vaccine Adjuvants (ed. Davies, G.) Methods Mol. Biol., 626, 117-130, Humana Press, Totowa, NJ, USA

Written by JohnG
Filed under: Cells