Erythrocyte fractionation by velocity sedimentation and discontinuous density gradient centrifugation

Abstract

Investigation of development, maturation and senescence in fish red blood cells depends on effective separation methods. Preliminary to initiation of studies of this type the erythrocytes of goldfish,Carassius auratus, and rainbow trout,Oncorhynchus mykiss, were fractionated by velocity sedimentation at unit gravity or discontinuous Percoll density gradient centrifugation in an attempt to separate and characterize enriched populations of juvenile and mature cells. Cytomorphological variables were assessed by microphotography (trout) and image analysis (goldfish). In both species velocity sedimentation led to fractions which were readily recognized in terms of mean length (maximum cell chord), area and shape (chord ratio, form factor), and sufficiently enriched for some types of study. Density gradient centrifugation also lead to cell separation. However, fractional densities appeared to reflect nuclear rather than cellular morphologies, and the fractions obtained were not readily categorized in relation to stage of development. Accordingly, at the present time, velocity sedimentation appears to offer the more suitable means for fractionating red cells for investigation of maturation and aging processes.