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.

Potential of endocervical lavage for cytogenetic investigation of fetal cells as determined by in situ hybridization.

VENTURINI, PIER LUIGI;FULCHERI, EZIO
1998-01-01

Abstract

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.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11567/254386
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