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(American Journal of Pathology. 2002;160:51-58.)
© 2002 American Society for Investigative Pathology

Short Communications

Enrichment, Immunomorphological, and Genetic Characterization of Fetal Cells Circulating in Maternal Blood

Giovanna Vona*, Christophe Béroud{dagger}, Alexandra Benachi{ddagger}, Alice Quenette{dagger}, Jean Paul Bonnefont§, Serge Romana, Yves Dumez{ddagger}, Bernard Lacour{dagger} and Patrizia Paterlini-Bréchot*{dagger}

From Unité INSERM 370,*
Faculté de Médecine Necker, Paris; and the Laboratoires de Biochimie A,{dagger}
de Génétique Médicale,§
and de Cytogenetique,
and the Service de Maternité,{ddagger}
Hôpital Necker-Enfant Malades, Paris, France

Fetal cells circulating in the peripheral blood of pregnant women are a potential target for noninvasive genetic analyses. They include epithelial (trophoblastic) cells, which are larger than peripheral blood leukocytes. We enriched circulating trophoblastic cells using the isolation by size of epithelial tumor cells (ISET) method. Peripheral blood was obtained at 11 to 12 weeks of pregnancy. Cells isolated by ISET were stained by hematoxylin and eosin or by immunohistochemistry. Large epithelial cells were microdissected and fetal cell identification was obtained by polymerase chain reaction with short tandem repeats and/or Y-specific primers. By analyzing only 2 ml of blood, we found a variable number (n = 1 to 7) of Y-positive cells (overall 15 of 23) in all of the six mothers carrying a male fetus. In contrast, none of the 26 cells isolated from seven mothers carrying a female fetus scored positive. Eleven cells were analyzed by using short tandem repeat-specific markers: six of them showed a fetal profile and five showed a maternal profile consistently with Y-specific results. Only one-fifth of the single cell DNA was used for fetal cell assessment, leaving enough material for further genetic tests. We also show that the ISET approach allows the performance of fluorescence in situ hybridization analyses and the detection of DNA point mutations in single microdissected cells. We conclude that this is a powerful approach to enrich circulating fetal cells and prove their fetal origin, and that it may have implications for noninvasive prenatal diagnosis of genetic disorders.


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