Potential of endocervical lavage for cytogenetic investigation of fetal cells as determined by in situ hybridization.
Aims: Transcervical samples following endocervical lavage from 28 women undergoing pregnancy termination were analysed in terms of fetal cells content and suitability for in situ hybridization analysis. A non fluorescent protocol of hybridization coupled to specific cytological procedures was used for simultaneous recognition and investigation on syncitiotrophoblast fragments. Methods: A series of endocervical washings was randomised to cytobrushing samples. Cervical washings were performed using variable volumes of physiologic saline solution (2ml, 5ml, 10ml). Single target in situ hybridization for chromosomes 1 and 18 was applied for determining a model of efficiency. Syncitial cells were recognized by means of cytoplasmic and nuclear stainings obtained with Eosin G (Diff-Quik) and haematoxylin respectively. Results: When 5 - 10ml of sterile solution were flushed multiple fragments of syncitio-trophoblast were almost constantly recovered. Conversely using 2ml of flushing solution no syncitia at all could be retrieved. Fetal cells were collected only in 50% of cervical cytobrushing. The per cell hybridization efficiency was on average 50%. Following in situ hybridization, a normal number of signal-products (2) was found in all nuclei analysed. Conclusion: During endocervical washing the amount of volume used is a critical factor to allow successful recovery of fetal cells. By using 5 - 10 ml of injecting solution, enough fetal material was generally obtained from each individual to guarantee the cytogenetic investigation for two chromosomes. The use of Diff-Quik and haematoxylin staining coupled to bright-field microscope facilitate the recognition of syncitia among a highly heterogeneous population of maternal cells. Further studies are ongoing to prospectively evaluate safety of the transcervical approach for sampling and analysis of fetal cells.
Bernardini
L
Department of OB/GYN, University Hospital of S. Martino, Genova, 16138, Italy. bernar01@aleph.it
Gerbaldo
D
Venturini
PL
Conte
N
Maritali
VA
Fulcheri
E