Fractionation and analysis of normal erythrocytes, sickle cells and reticulocytes; removal of erythrocytes from blood and bone marrow

A number of density gradient strategies have been developed for the fractionation of human erythrocytes according to their age. As the cells age, so their density tends to increase; reticulocytes therefore have the lowest densities. 

Reticulocytes have frequently been partially purified on discontinuous gradients of arabinogalactan; the actual density range being quite variable, sometimes broad, e.g. 1.066-1.220 g/ml [1], sometimes narrow, e.g. 1.083-1.124 g/ml [2]. 

These gradients have entailed the use of unusually high RCFs of 27,000g [1] 74,000g [2] for approx 1 h. Such high RCFs are rarely used for other types of cell and are often detrimental to cell function; the viscosity of arabinogalactan gradients maybe a contributory factor. 

Continuous gradients of metrizamide (12%- 27%) equivalent to a density range of approx 1.07-1.15 g/ml were used by Lodish et al [3] to purify rabbit reticulocytes and again a relatively high RCF was used – approx 27,000 g for 1 h.