title: Fetal cell isolation from maternal blood cultures by flow cytometric hemoglobin profiles. Results of a preliminary clinical trial.
abstract: OBJECTIVE
  We conducted a trial to test if the blood of pregnant women contains fetal clonogenic erythroid cells the progeny of which can be identified and isolated by a newly developed flow-sorting procedure.
  
  
  METHODS
  We have previously demonstrated the identification of fetal nucleated red cells in cocultures of fetal and adult blood. The procedure is based on profiles of the correlated contents of fetal and adult hemoglobin (HbF and HbA, respectively), using antibodies specific for the different hemoglobin chains. In such profiles, fetal cells contain only HbF, while the vast majority of adult cells contain either only HbA or a combination of HbA and HbF. HbF+ HbA- cells are flow sorted and fetal cells identified by fluorescence in situ hybridization, using chromosome-specific probes. This technique provides a yield that approaches 100%, meaning that fetal cells will be found even if the culture contains only a single fetal erythroid colony among thousands of maternal colonies. Peripheral blood samples were obtained from 11 women carrying chromosomally normal male fetuses, from 5 women carrying trisomy 21 fetuses, and from 2 women carrying trisomy 18 fetuses. A further six samples came from women with an unknown fetal karyotype. As positive controls, we used blood samples drawn after termination procedures that tended to induce some fetomaternal hemorrhage. In parallel to the method being tested, we employed alternative techniques of fetal cell detection: one third of the mononuclear cell preparations from each maternal blood sample was not cultured but labeled with anti-HbF antibodies for flow sorting of F+ cells. Ten percent of the total harvested cell population of each culture was subjected to quantitative polymerase chain reaction analysis targeting a Y-chromosome-specific sequence.
  
  
  RESULTS
  Most post-termination blood samples yielded fetal cells with high purity which demonstrates the validity of the method. However, no fetal cells were found in any of the maternal blood samples with normal or abnormal pregnancies, neither before nor after culture.
  
  
  CONCLUSIONS
  We conclude that a cell culture approach targeting clonogenic erythroid cells offers no advantage over established methods of direct isolation.
authors.affiliation: Division of Genetics, Department of Pediatrics, New England Medical Center and Tufts University Medical School, Boston, MA 02111, USA. rbohmer@lifespan.org
authors.forename: Ralph M
authors.initials: RM
authors.lastname: Bohmer
authors.forename: Helene P
authors.initials: HP
authors.lastname: Stroh
authors.forename: Kirby L
authors.initials: KL
authors.lastname: Johnson
authors.forename: Erik S
authors.initials: ES
authors.lastname: LeShane
authors.forename: Diana W
authors.initials: DW
authors.lastname: Bianchi
mesh_terms: D018929:Cell Culture Techniques; D002469:Cell Separation / Q000379:methods*; D002887:Chromosomes, Human, Pair 18; D004314:Down Syndrome / Q000097:blood; D015672:Erythroid Precursor Cells / Q000166:cytology*; D005260:Female; D005312:Fetal Blood / Q000166:cytology*; D005319:Fetal Hemoglobin / Q000032:analysis*; D005434:Flow Cytometry; D005865:Gestational Age; D006441:Hemoglobin A / Q000032:analysis*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008297:Male; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D014314:Trisomy
chemical_list: D006441:Hemoglobin A; D005319:Fetal Hemoglobin