Cytogenetical diagnosis in paraf®n-embedded fetoplacental tissue using comparative genomic hybridization Tulin Ozcan*, Nicole Burki, Vinita Parkash, Xiaohua Huang, Tanja Pejovic, Maurice J. Mahoney and David C. Ward Department of Genetics, Yale University School of Medicine, New Haven, CT, USA Department of Pathology, Yale University School of Medicine, New Haven, CT, USA Department of Obstetrics and Gynecology, Yale University School of Medicine, New Haven, CT, USA Comparative genomic hybridization (CGH) is a FISH-related technique used to assess global chromosomal aberrations in a variety of human tumours. Recently CGH has been applied to cytogenetic analysis of fresh frozen fetoplacental tissues. Here we report the application of CGH to paraf®n-embedded placental samples. Ten samples from paraf®n-embedded blocks of 6 control placentas and fetoplacental tissue from 10 aneuploidies, and 2 unbalanced aberrations were evaluated. Balanced karyotype pro®les were obtained from samples of healthy placentas and all samples from the same placenta appeared to have similar con®dence intervals. CGH analysis of four cases of trisomy 21, three cases of trisomy 18, one case of trisomy 13, one case of trisomy 15 and one case of trisomy 7 all showed overrepresentation of the respective trisomic chromosome. The CGH pro®le was also in accordance with the karyotyping of a case with isochromosome 21. The CGH pro®le of a case with der (2)t(2;6)(q37.3;q22.2) revealed partial trisomy for chromosome 6 between q21 and q27. CGH may be a useful adjunct in prenatal genetic diagnosis when retrospective diagnosis is needed from archival samples. Copyright # 2000 John Wiley & Sons, Ltd. KEY WORDS: comparative genomic hybridization; aneuploidy; fetal karyotype INTRODUCTION In prenatal diagnosis, conventional Giemsa banding of metaphase chromosomes and ¯uorescent in situ hybridization (FISH) are the most widely used techniques for detecting genetic aberrations. While FISH can be applied in situations where only interphase cells are available, with interphase FISH, only a few regions of the genome can be explored at a time necessitating a prediction of expected cytogenetic abnormalities based on phenotype. Comparative genomic hybridization (CGH) is a molecular cytogenetic approach using ratio-imaging FISH for detection of unbalanced chromosomal aberrations (Kallioniemi et al., 1992). It has mostly been used as a research tool for evaluation of human solid tumours where access to metaphase spreads is often limited. The technique uses DNA extracted directly from the tissue to be analysed and therefore circumvents the need for metaphase spreads and cell culture. A mixed probe of labelled test genomic DNA and an equimolar amount of differently labelled reference DNA prepared from cells with normal chromosome complements is used for chromosomal in situ suppression hybridization to normal metaphase spreads. Hybridized test and control DNA sequences are detected with different ¯uorochromes. The ratios of ¯uorescence intensities re¯ect the relative copy number in the test genome compared with the control genome. Thus, CGH provides the possibility for searching the whole genome for unbalanced genetic material in a single hybridization reaction. The use of CGH for fresh frozen fetoplacental samples has been reported in a few studies (Bryndorf et al., 1995; Daniely et al., 1998;1999; Yu et al., 1997). However, the applicability of this technique in paraf®n- embedded fetoplacental tissue has not been evaluated. A policy of genetic analysis is recommended for recurrent abortions and all stillbirths to avoid parental anxiety, unnecessary diagnostic tests and to provide appropriate counselling of the parents for future pregnancies (Wolf and Horger 1995; ACOG Commit- tee Opinion No. 178, 1996). Work-up for recurrent pregnancy losses is usually started after two or three losses from which fresh fetoplacental tissue is no longer available for culture. CGH may be a potentially powerful method for detection of prenatal cytogenetic aberrations when retrospective karyotyping is needed, i.e. in cases of recurrent abortion and fetal demise. The purpose of this study was to evaluate the applicability of CGH in paraf®n-embedded feto- placental samples for genetic analysis. MATERIAL AND METHODS Tissue from paraf®n-embedded blocks of six term placentas and 12 samples of abortion material diagnosed with aneuploidies or unbalanced aberra- tions by conventional karyotyping were evaluated by CGH. A total of 10 biopsies from chorionic villi of six healthy term pregnancies collected prospectively were classi®ed as the normal group. Though conventional karyotyping of these six cases was not done, they were *Correspondence to: T. Ozcan, Yale University School of Medicine, Department of Genetics, 333 Cedar Street, SHM I-147, New Haven CT 6510, USA. E-mail: tulin.ozcan@yale.edu PRENATAL DIAGNOSIS Prenat Diagn 2000; 20: 41±44. CCC 0197-3851/2000/010041±04$17.50 Copyright # 2000 John Wiley & Sons, Ltd. Received: 15 June 1999 Revised: 30 September 1999 Accepted: 8 October 1999