Transcervical recovery of fetal cells from the lower uterine pole: reliability of recovery and histological/immunocytochemical analysis of recovered cell populations.
The aim of this work was to isolate, enumerate and attempt the identification of fetal cells recovered from the lower uterine pole. Immediately before elective termination of pregnancy at 7-17 weeks gestation, samples were recovered by transcervical flushing of the lower uterine pole (n = 108) or transcervical aspiration of mucus from just above the internal os (n = 187), and their contents examined using histological, immunohistochemical and molecular techniques. Syncytiotrophoblasts were identified morphologically in 28 out of 89 (31%) and 50 out of 180 (28%) flushings and aspirates respectively (mean 29%). Immunocytochemistry with monoclonal antibodies (mAbs) recognizing trophoblast or epithelial cell antigens on a smaller number of samples (n = 69) identified putative placental cells in 13 out of 19 (68%) and 25 out of 50 (50%) flushings and aspirates respectively (mean 55%). These included groups of distinctive cells with a small, round, hyperchromatic nucleus, strongly reactive with mAbs PLAP, NDOG1 and FT1.41.1. Smaller groups of larger, amorphous cells, usually containing multiple large, pale staining nuclei, reactive with mAb 340 and to a lesser degree with mAb NDOG5 were also observed. Taking cellular morphology and immunophenotype into consideration, the smaller uninucleate cells were likely to be villous mesenchymal cells, while the larger cells were possibly degrading villous syncytiotrophoblast. There was no significant difference in the frequency of fetal cells obtained by the two recovery methods. Squamous or columnar epithelial cells, labelled strongly with antibodies to cytokeratins or human milk fat globule protein, were observed in 97% (29 out of 30) of aspirates. The use of cervagem in a small number of patients prior to termination of pregnancy did not appear to influence the subsequent recovery of placental cells. Y-specific DNA was detected by polymerase chain reaction (PCR) in 13 out of 26 (50%) flushings and (99 out of 154) 64% aspirates analysed (mean 62%). In-situ hybridization (ISH) revealed Y-specific targets in 40 out of 69 (60%) of aspirates analysed. A comparison of PCR data obtained from transcervical recovered samples and placental tissues showed a concordance of 80% (76 out of 95), with 10 false positives. Comparing the PCR data from tissues with data derived by ISH from 41 aspirates gave a concordance of 90% with two false positives. Although syncytiotrophoblasts were much more likely to be present in samples containing immunoreactive placental cells, the detection rates of fetal-derived DNA were similar regardless of the morphological and/or immunological presence of placental cells. We conclude that the transcervical recovery of fetal cells, while promising, requires considerable additional effort being expended in further research and development, particular in the sampling procedure.
Miller
D
D
Centre for Reproduction, Growth and Development, University of Leeds, UK.
Briggs
J
J
Rahman
M S
MS
Griffith-Jones
M
M
Rane
V
V
Everett
M
M
Lilford
R J
RJ
Bulmer
J N
JN
D000911:Antibodies, Monoclonal; D002584:Cervix Uteri; D005260:Female; D005333:Fetus / Q000166:cytology*; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D017403:In Situ Hybridization; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D012732:Sex Determination Analysis; D013048:Specimen Handling / Q000379:methods*; D013396:Suction; D007507:Therapeutic Irrigation; D014599:Uterus
D000911:Antibodies, Monoclonal