Identification of fetal nucleated red cells in co-cultures from fetal and adult peripheral blood: differential effects of serum on fetal and adult erythropoiesis.
Seeking to optimize a novel method of isolating rare fetal erythroid cells in cultures from maternal blood, we have explored the effects of serum supplement on fetal and adult erythropoiesis. We used flow cytometry and sorting after labelling with antibodies to fetal haemoglobin (HbF) and adult haemoglobin (HbA). In adult blood-derived cultures, most nucleated red cells accumulated either only adult haemoglobin (F-A+) or a combination of fetal and adult haemoglobin (F+A+). Only a few were F+A-. Serum affected the proportions of adult cells expressing fetal haemoglobin (both F+A- and F+A+), which were minimized, but not eliminated altogether, with the use of charcoal-treated sera at low concentrations. In contrast, the expansion of fetal red cells, which made only fetal haemoglobin (F+A-) during at least one week of culture, was strongly increased with the use of charcoal treated sera, due to the removal of a charcoal-absorbable inhibitor. In co-cultures of fetal and adult erythroid cells, fetal cells could be enriched in the order of 200-fold by flow sorting with the F+A- criterion. However, since adult F+A- cells could not be suppressed completely, the purity of sorted fetal cells still depended on the relative numbers of fetal and maternal erythroid clonogenic cells in the blood sample. Thus, we demonstrate a method by which fetal nucleated red cells potentially present in maternal blood cultures can be identified and isolated from the vast majority of maternal erythroid cells, based on their correlated contents of fetal and adult haemoglobin.
Division of Genetics, Department of Pediatrics, New England Medical Center and Tufts University Medical School, Boston, MA 02111, USA. ralph.bohmer@es.nemc.org
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D000328:Adult; D001769:Blood; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D002606:Charcoal; D018920:Coculture Techniques; D004912:Erythrocytes / Q000502:physiology* / Q000648:ultrastructure*; D015672:Erythroid Precursor Cells / Q000166:cytology; D004920:Erythropoiesis; D005260:Female; D005312:Fetal Blood / Q000166:cytology*; D005319:Fetal Hemoglobin / Q000032:analysis; D005434:Flow Cytometry; D006441:Hemoglobin A / Q000032:analysis; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D011247:Pregnancy
D002606:Charcoal; D006441:Hemoglobin A; D005319:Fetal Hemoglobin