Identification of CD71 (transferrin receptor) expressing erythrocytes by multiparameter-flow-cytometry (MP-FCM): correlation to the quantitation of reticulocytes as determined by conventional microscopy and by MP-FCM using a RNA-staining dye

Abstract

Reticulocytes express the CD71-defined antigen, the transferrin receptor. This report describes how by means of dual-colour immunofluorescence using MP-FCM (multiparameter-flow-cytometry) CD71+ erythrocytes can be detected regularly in blood of healthy adults. Percentages of these CD71+ erythrocytes were compared to the percentages of reticulocytes as determined by conventional microscopy using brilliant cressyl blue, and to the percentages of erythrocytes with high RNA content, as detected by MP-FCM using a RNA-staining dye (thiazole-orange). Only about two-thirds of the percentages determined by the two latter methods were detected by MP-FCM using the CD71 expression for definition of reticulocytes. Studying clinical samples, however, including both specimens with very low and very high numbers of reticulocytes, almost identical percentages were determined by all the three methods described. Studying reticulocytes in vitro, a rapid decline of the expression of the transferrin receptor was observed on reticulocytes. Due to the differential expression of the transferrin receptor on reticulocytes, different subsets of reticulocytes could be identified. The dual-colour MP-FCM method described allows for the characterization and enumeration of immature erythrocytes, representing the major subset of reticulocytes as determined by conventional methods. Furthermore, it allows for subset dissection of reticulocytes.