Chorionic villus culture for prenatal diagnosis of chromosome defects: Reduction of the long‐term cultivation time

Abstract

We report in detail two series of chorionic villus cultivation for prenatal chromosomal diagnosis. Chorionic villi were sampled from both first- and second-trimester pregnancies. One hundred cultures were treated with trypsin-EDTA for 2 h and collagenase overnight, (method A) and 100 were treated with trypsin-EDTA for 1 h and collagenase for 2 h (method B). Using short-term enzymatic digestion, the cultivation time was reduced from 14 days to 6 days. Sufficient amounts of metaphases of good quality were present in 93 per cent of primary cultures harvested in situ, whereas enough metaphases of sufficiently good quality were in most cases present only after subcultivation of the cultures using method A. The decrease in cultivation time obtained is probably due to a higher yield of viable cells in monocellular suspension, an increased attachment efficiency, and a more rapid attachment of single cells (within 24 h).

Accuracy of abnormal karyotypes after the analysis of both short- and long-term culture of chorionic villi

“…The very low percentage of MCC in the present study (0.4%) is not only the result of careful dissection of the material, the culture method and the choice of appropriate medium (Saura et al, 1997;Smidt-Jensen et al, 1989), but is also due to the fact that all LTCvilli are harvested in situ, in our laboratory. There were no diagnostic errors due to MCC.…”

“…For the STC-villi preparation 15 mg were incubated overnight using¯uorodeoxyuridine (FdU) synchronization (Gibas et al, 1987). The remaining villi were simultaneously used for the preparation of LTC-villi, which consisted of a trypsin-EDTA and collagenase treatment at 37uC for 60 and 120 min, respectively (Smidt-Jensen et al, 1989). Cells were cultured in Lab-tek1II Chamber slides 2 (Nalge Nunc International, Naperville, IL, USA) and harvested in situ after 5±7 days.…”

Cytogenetic confirmation of a positive NIPT result: evidence-based choice between chorionic villus sampling and amniocentesis depending on chromosome aberration

“…The cytotrophoblast originates from the trophoblast of the embryo and traditionally is studied in direct or short-term-cultured villi (STC-villi) [15] and nowadays is tested with NIPT [10]. In contrast, the mesenchymal core of CV, which conventionally is studied in long-term-cultured villi (LTC-villi) [16], better reflects the fetal karyotype. This is due to its embryonic origin, which is the same as that of the fetus itself (inner cell mass) [17,18].…”

“…For the other trisomies that often are found in CV cytotrophoblast (3,7,8,9,15,16,20,22), the numbers in the ACC CVS database are very small and only individual case reports are published. Therefore, no reliable risk figure as for trisomy 13, 18, or 21 can be drawn from the database or literature.…”

Prevalence of Tetraploid Metaphases in Semidirect and Cultured Chorionic Villi

“…STC-and LTC-villi slides were prepared according to standard techniques [18,19]. The LTC slides were harvested in situ after 5-7 days of culturing.…”