Abstract for presentation at 11th International Congress of Human Genetics

Quality aspects in prenatal cytogenetic diagnostics: Effect of various factors on the quality of in situ culture and slide preparation of primary amniotic fluid and chorionic villi biopsy cells

  • Birgit Sikkema-Raddatz, Department of Clinical Genetics, University Medical Center Groningen, The Netherlands
  • Ron Suijkerbuijk, Department of Clinical Genetics, University Medical Center Groningen, The Netherlands
  • Katelijne Bouman, Department of Clinical Genetics, University Medical Center Groningen, The Netherlands
  • Bauke de Jong, Department of Clinical Genetics, University Medical Center Groningen, The Netherlands
  • Charles Buys, Department of Clinical Genetics, University Medical Center Groningen, The Netherlands
  • Gerard te Meerman, Department of Clinical Genetics, University Medical Center Groningen, The Netherlands

    • Purpose: To investigate the effect of various factors on cell culturing and slide preparation from cells from amniotic fluid (AF) and chorionic villus biopsies (CVB).
    Methods: From 579 cases, received in a one-year period, we retrospectively investigated the effects of volume and appearance of the admitted AF specimen, the gynaecologist performing the amniocentesis, the reason for referral, the week of gestation and the culture medium onto. In addition, we tested the effect of decreased oxygen (O2) levels during culturing of 30 new AF cases. As a quality measure for cell culture, the total number of colonies on day 5-6 and the total culture time until slide preparation were determined. During slide preparation, controlled experimental variation in the composition of fixative, relative humidity, ambient temperature and air flow of primary CVB and AF-cell cultures was introduced. As a quality measure, the degree of spreading of metaphases and the quantity of cytoplasm present were assessed. For AF cells, 1371 metaphases were screened, for CVB cells 1391 metaphases. All factors were evaluated by analysis of regression or variance.
    Results: Amongst the parameters tested, only the variation in appearance of the AF and O2 tension had a rather major impact. Bloody or brown AF resulted in an extended culture time of 3 days more on an average of 8-9 days, while a 5% O2 tension reduced culture time for at least 1-2 days. Regarding slide preparation of AF and CVB cells, chromosome spreading was mostly (but moderately) affected by the composition of fixative. As for the presence of cytoplasm in metaphase spreads, for AF cells this was mostly influenced by the relative humidity and for CVB cells by air flow.
    Conclusion: In general, our prenatal cytogenetic practice has proven to be robust and reliable. Most conditions examined appeared not as critical within the range of variation. Therefore, expensive measures for quality-controlling of these conditions are not required.

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