Human peripheral blood and bone marrow cell separation using density gradient centrifugation on Lymphoprep and Percoll in haematological diseases

Abstract

Density gradient centrifugation on Lymphoprep as described by Böyum [1] was used for mononuclear cell separation of peripheral blood (PB) and bone marrow (BM) samples in healthy subjects and in haematological diseases. The cells were analysed morphologically from cytocentrifuge preparations stained with May-Grünwald-Giemsa. At least 200 cells per preparation were counted manually. The separation results of PB samples in healthy subjects were comparable with those described by Böyum. This method was also suitable for the separation of lymphocytes and blast cells in chronic lymphoproliferative diseases and in acute leukaemias, respectively. In other haematological diseases, however, the mononuclear cell fractions of PB samples and especially of BM samples were contaminated with myeloid and erythroid cells. Percoll gradients were used with success for the enrichment of myeloid BM cells and PB reticulocytes. However, absolutely pure myeloid cell fractions from different maturation stages could not be separated. The results indicate that cells to be investigated should be morphologically examined, to avoid erroneous interpretation of biochemical and functional activity of the cells.

Key words:

• blood cells
• cell separation
• density gradient
• haematological diseases