Density Gradient Centrifugation

The enrichment of cells and the optimisation of cell separation methods play a decisive role for many researchers. Various approaches exist for sorting different cell types, with the choice of the appropriate method often depending on the cell type or cell subset. One of the faster and more cost-effective methods for separating cells is density gradient centrifugation. In our article"Cell separation and cell separation methods" we have summarised the various options for enriching cells from heterogeneous mixtures, such as blood.

Based on the different densities of the individual blood components, these can be separated by single-stage centrifugation with the appropriate density gradient medium. The technique is based on the fact that particles with different densities have different sedimentation rates during centrifugation. Heavy particles such as erythrocytes sediment quickly, while lighter cells such as thrombocytes sediment more slowly. The following figure shows the distribution of the most important cell populations in blood. Erythrocytes have the highest density and platelets the lowest density.

The following figure (Figure 1) shows the distribution of blood cells depending on their density. Thrombocytes (1) have the lowest density, erythrocytes (2) the highest density. The number of cells per population is also shown. Erythrocytes represent the largest population, platelets the second largest population.

Figure 1: Illustration of the cell populations of blood as a function of density  

The cell populations thrombocytes (1), monocytes (2), lymphocytes (3), basophil granulocytes (4), neutrophil granulocytes (5), eosinophil granulocytes (6) and erythrocytes (7) differ in density and cell number. The different densities of the cell populations are used for enrichment with the aid of density gradient media.

As can be seen from the figure and the table, the density of the cells or cell populations is a distribution around a mean value. In addition, the densities of the different cell populations overlap. The range as well as the distribution around the mean value are not fixed values. Fluctuations in density can be influenced by various factors. One of the most important factors influencing the density of the cells is the hydration of the cells. Another important factor influencing the density of whole blood cells is the time outside the donor organism.

Figure 2: Distribution of layers after density gradient centrifugation of whole blood with PBMC spin medium. PBMC (2) collect between plasma (1) and density medium (3). Erythrocytes, granulocytes and dead cells pass through the medium and are concentrated at the bottom of the tube (4).