[First attempts of detecting fetal cells in the maternal circulation].
BACKGROUND
In prenatal diagnosis there is great interest for noninvasive diagnostic methods. Authors report their first results in detecting fetal cells in the maternal circulation during pregnancy.


OBJECTIVE
The aim of the study was to detect fetal gender from maternal peripheral blood samples during pregnancy.


METHODS
Authors have analysed fetal nucleated red blood cells. In 12 cases after a double density Percoll gradient separation they labelled the surface antigens of the cells with anti-glycophorin-A and anti-CD45 fluorescent antibodies, did an intracellular staining of the epsilon haemoglobin chain, and analysed the cells with flow cytometry. The CD45 negative/glycophorin-A positive/epsilon-haemoglobin chain positive cells were considered as fetal cells. Having the results, in another 13 cases magnetic activated cell sorting with CD71 antibody were used as an enrichment step. Authors made an intracellular staining of the epsilon haemoglobin chain, the positive cells were isolated by micromanipulation, and analysed by single cell fluorescent polymerase chain reaction. Primers for the amelogenin gene were used to detect fetal gender.


RESULTS
Only the Percoll enrichment step itself is not enough for using the samples for diagnostic molecular-biologic examinations, a following enrichment step is needed. For this the authors used magnetic activated cell sorting with CD71 antibody. With the help of this enrichment step, after the intracellular staining of the epsilon haemoglobin chain the direct micromanipulator isolation of the epsilon haemoglobin chain positive cells could be done. After analysing single cells by fluorescent polymerase chain reaction, in 8 out of the 11 comparable cases the results were similar to those, what was found during the genetic amniocentesis. In 2 cases from this 8, genetic amniocentesis proved Klinefelter syndrome, which they could also confirm with the examination of fetal cells in the maternal circulation.


CONCLUSIONS
The results of the study suggest that the method described above can be useful in prenatal genetic diagnosis, and improving it could be useful to detect other genetic abnormalities (chromosomal abnormalities, single gene disorders) as well.
Nagy
Gyula Richárd
GR
Semmelweis Egyetem, Altalános Orvostudományi Kar, I. Szülészeti és Nogyógyászati Klinika, Budapest. nagygyr@noi1.sote.hu
Bán
Zoltán
Z
Sipos
Ferenc
F
Fent
János
J
Oroszné Nagy
Judit
J
Beke
Artúr
A
Furész
József
J
Papp
Zoltán
Z
D015703:Antigens, CD; D000944:Antigens, Differentiation, B-Lymphocyte; D002469:Cell Separation / Q000379:methods*; D004900:Erythroblasts; D005260:Female; D005312:Fetal Blood; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D005914:Globins / Q000032:analysis; D006801:Humans; D007713:Klinefelter Syndrome / Q000175:diagnosis / Q000235:genetics; D008280:Magnetics; D016133:Polymerase Chain Reaction; D011205:Povidone; D011247:Pregnancy; D011296:Prenatal Diagnosis; D011990:Receptors, Transferrin; D019849:Sex Determination Processes; D012822:Silicon Dioxide
D015703:Antigens, CD; D000944:Antigens, Differentiation, B-Lymphocyte; C068126:CD71 antigen; D011990:Receptors, Transferrin; C016039:Percoll; D012822:Silicon Dioxide; D005914:Globins; D011205:Povidone