Separation and Size Distribution of Red Blood Cells of Diverse Size, Shape, and Origin by Flow/Hyperlayer Field-Flow Fractionation

Red blood cells (RBCs) from human, equine, canine, feline, and bovine whole blood samples have been separated and characterized by high-speed flow/hyperlayer field-flow fractionation (Fl/HyFFF). The elution-based separation of RBCs by this method is based mainly on the size and shape of the cell particles. The typical separation time for RBCs is less than 3 min. Size distributions can be derived from the fractograms of cell samples using a calibration plot based on retention data for uniform polystyrene beads. The method is shown to be effective both to separate and to characterize cell populations, including those with cells of abnormal shape and size. In order to investigate differences in cell morphology, shape and size changes induced by 500,000 Da Dextran were monitored. The changes in the Fl/HyFFF elution profiles indicate that the RBCs decrease in size but become partially aggregated in the presence of the dextran. These changes were found to depend on polymer concentration and specific blood samples. Some of the results from Fl/HyFFF were compared with those from the Coulter counter and from microscopy.