Detection and relocation of cord blood nucleated red blood cells by laser scanning cytometry

• Published in: Cytometry (New York, N.Y.) N.Y.), 2002-06, Vol.48 (2), p.87-92
• Main Authors: Hennerbichler, Simone, Schmied, Rudolf, Petek, Erwin, Kroisel, Peter M., Pertl, Barbara, Tiran, Beate, Dohr, Gottfried, Wintersteiger, Reinhold, Sedlmayr, Peter
• Format: Article
• Language: English
• Subjects: Cell Nucleus - metabolism; Chromosomes, Human, Y; erythroblasts; Erythroblasts - cytology; Female; Fetal Blood - cytology; Humans; Image Cytometry - methods; In Situ Hybridization, Fluorescence; Infant, Newborn; LSC; Male; Microscopy, Confocal; Prenatal Diagnosis; prenatal diagnostics; Staining and Labeling
• ISSN: 0196-4763; 1097-0320
• DOI: 10.1002/cyto.10112

Background Fetal nucleated red blood cells (NRBC) present in the peripheral blood of pregnant women at low frequency are a potential target for noninvasive prenatal diagnostics. Methods CD71‐enriched cells from male cord blood (CB) were stained for the gamma chain of HbF (Hb‐γ) and cytocentrifuged. Fluorescence in situ hybridization (FISH) was done for the Y chromosome. Following staining of the nucleus with TO‐PRO‐3, laser scanning cytometry was performed. Artificial mixtures of small volumes of male CB and blood drawn from nonpregnant females were analyzed. Results In CB, 59% of events double positive for Hb‐γ and TO‐PRO‐3 were identified as CB‐NRBC. In contamination studies, male fetal CB‐NRBC were identified perfectly on the basis of morphologic characteristics and FISH reactivity following relocation and visual assessment. Mean recovery was 8.7%. Conclusions Laser scanning cytometry of preenriched fetal NRBC may offer a promising way for noninvasive prenatal diagnostics. This is because it provides a virtual enrichment step and the position on the slides of cells visually confirmed to correspond to fetal NRBC is known. Further experimental procedures on well‐defined and located target cells may be feasible. Cytometry 48:87–92, 2002. © 2002 Wiley‐Liss, Inc.