Comparison of two techniques for transcervical cell sampling performed in the same study population.
OBJECTIVE
The aim of this study was to evaluate and compare the presence of fetal cells in transcervical cell (TCC) samples collected in the first trimester of pregnancy by two different procedures [mucus collection and intrauterine lavage (IUL)], performed consecutively in the same subjects scheduled for elective termination of pregnancy (TOP).


METHODS
A total of 126 mucus/IUL sample pairs were retrieved from pregnant women immediately before TOP at a gestational age ranging from 7 to 12 weeks; at termination, samples of placental tissue were collected in all cases. All mucus samples were analysed by a polymerase chain reaction (PCR) assay and, in a subset of experiments involving 56 specimens, also by fluorescence in situ hybridization (FISH) procedure. IULs were divided in two aliquots, one for PCR analysis and one for the preparation of FISH slides. All placental tissue samples obtained at termination were analysed by FISH for fetal sexing. The PCR assay for fetal sex determination was performed by using, in a multiplex reaction, primers for SRY (Y chromosome sex-determining region, 738 bp) and HUMARA (human androgen receptor on the X chromosome, 280 bp) genes. The FISH analysis was carried out using direct-labelled commercial probes for X chromosome alpha-satellite (DXZ1, Xp11.1-q11.1, spectrum green) and Y chromosome alpha-satellite (DYZ3, Yp11.1-q11.1, spectrum orange) regions.


RESULTS
In samples from known male pregnancies (n = 67), full concordance between IUL and mucus results could be found in 11 cases (16.4%); in 41 cases, Y chromosome material was detected by FISH (n = 2), by PCR (n = 5) or both (n = 34) in IUL samples, but not in the corresponding mucus samples. Y chromosome material was not documented in 10 mucus/IUL sample pairs. In 5 cases, the FISH (n = 2), the PCR (n = 1) or both (n = 2) failed to detect Y chromosome material in IULs, which was detected, however, by PCR in the corresponding mucus samples. Overall, correct sex prediction was achieved in 55/67 IULs (82%) and in 16/67 (23.9%) mucus samples from male pregnancies. In samples from known female pregnancies (n = 56), full concordance between results of IUL/mucus pairs and those on placental samples could be found in 53 cases (94.6%); in 3 cases, Y chromosome material was documented by PCR in mucus samples, but not in the corresponding IULs. Correct sex prediction was therefore achieved in 56/56 IULs (100%) and in 53/56 (94.6%) mucus samples from female pregnancies.


CONCLUSIONS
This study provides evidence that, among TCC sampling techniques, IUL, but not mucus collection, can yield fetal cells in a constant and reliable fashion, which is a basic prerequisite for possible clinical usage. This suggestion had already emerged from some previous investigations but, owing to the study design, differences in study populations can no longer be used to explain the very different and sometimes-conflicting results reported in earlier studies.
Cioni
Riccardo
R
Department of Gynecology, Perinatology and Human Reproduction, University of Florence, Viale Morgagni, Florence, Italy. rcioni@yahoo.it
Bussani
Cecilia
C
Scarselli
Benedetta
B
Bucciantini
Sandra
S
Marchionni
Mauro
M
Scarselli
Gianfranco
G
D000028:Abortion, Induced; D003581:Cytodiagnosis / Q000379:methods*; D003584:Cytological Techniques / Q000379:methods*; D003944:Diagnostic Techniques, Obstetrical and Gynecological; D005260:Female; D005315:Fetal Diseases / Q000175:diagnosis*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods; D016133:Polymerase Chain Reaction / Q000379:methods; D011247:Pregnancy; D011261:Pregnancy Trimester, First; D007507:Therapeutic Irrigation / Q000379:methods; D014599:Uterus