Density fractionation of erythrocytes by Percoll/hypaque results in only a slight enrichment for aged cells.
Density fractionation has served for many years as a standard procedure for the isolation of aged erythrocyte populations; however, a quantitative evaluation of the technique has not been available. This report describes such an analysis. Rabbits were infused intravenously with N-hydroxysuccinimido biotin to biotinylate greater than 90% of all circulating erythrocytes; the enumeration of these biotinylated cells was possible by monitoring their ability to bind avidin-coated microspheres. The biotinylated erythrocytes were shown to have a normal in vivo survival. As a result of the normal senescence process, only aged red cells would have membrane-bound biotin 50 days after biotinylation. At this time, the rabbit red cells were fractionated over Percoll/hypaque to determine the ability of density fractionation to enrich for the aged, biotinylated cells. Density fractionation resulted in an average enrichment for aged cells of only 2-3-fold above that present in a random population.
Dale
G L
GL
Department Molecular and Experimental Medicine, Research Institute of Scripps Clinic, La Jolla, CA 92037.
Norenberg
S L
SL
D000818:Animals; D001710:Biotin / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D004905:Erythrocyte Aging; D004912:Erythrocytes / Q000378:metabolism; D011205:Povidone; D011817:Rabbits; D015203:Reproducibility of Results; D012822:Silicon Dioxide
C016039:Percoll; D001710:Biotin; D012822:Silicon Dioxide; D011205:Povidone