Repetitive probes have proved useful for the enum eration of chromosomes 18, X and Y in sam ples of uncultured amniotic fluid (Guyot et al., 1988; Christensen et al., 1992; Cacheux et al., 1994).
In the present investigations the use of repetitive probes for chromosomes X and Y correctly d eterm in ed the fetal sex in all 127 cases ev alu ated including one Klinefelter fetus in which 47 out of 50 nuclei (94%) displayed two X hybridisation sig n a ls a n d one Y h y b rid is a tio n signal. Signals c o rre sp o n d in g to the Y chrom osom e w ere never seen in female fetuses and only 14 female cells in total out of 2798 show ed one instead of the expected two signals corresponding to the X chromosome.
The use of p robe D18Z1 (Oncor) w as rep o rted by Cacheux et al (1994). They rep o rted th at 97% of cells in norm al sam ples of am niotic fluid displayed two hybridisation signals w ith no norm al cells displaying three signals. Using this probe they correctly identified two cases of trisom y 18. In 87% and 84% of the nuclei in these two cases they observed three hybridisation spots. In the present series of 27 sam ples investigated using the probe D18Z1 there was one case of trisomy 18. In this case ( 727/97) 78% of the nuclei displayed three hybridisation spots. Cacheux et al (1994) reported a hybridisation efficiency of 99% and never observed three signals in disom ic samples. The 26 norm al samples in this series show ed only tw o nuclei w ith three signals (cases nos 1958/93 an d 1948/93). These cells accounted for only 2% and 1.3% respectively of the total num bers of
signals scored from each case. The hybridisation efficiency ranged from 72-100% and of the 27 samples 16 (59%) show ed a hybridisation efficiency of at least 98%. Cacheux et al (1994) co n clu d ed th a t the p ro b e D 13Z1/D 21Z1 (O ncor) w as unreliable for prenatal detection of trisomies 13 and 21 in uncultured amniocytes. The use of this probe and other 13/21 alpha satellite DNA probes hybridising to both chromosomes 13 and 21 were investigated in the current study. Resolving four signals in norm al cells and five signals in cells w ith either trisom y 21 or trisomy 13 was found to be less sensitive than resolving two as opposed to three signals. In ad d itio n variations in alphoid D N A sequences have lead to false results w ith this probe (Mizimoe and Young, 1992; Verma and Luke, 1992), and this has b een observed in the cu rren t stu d y u sin g m etap h ase sp re ad s from laboratory personnel. A furthur reason for not using using the cross hybridising 13/21 centrom eric probe is that Robertsonian translocation trisom ies w ould be undetected.
In the present study three other centromeric probes produced in the laboratory have been u sed on in terp h ase cells in diagnostic sam ples. These are probes specific for chrom osom es 9 (cCMP9.27), 16 (pSE16) and 18 (L1.84). All three probes have been show n to be capable of distinguishing betw een disom ic and trisomie cells in interphase and have the potential to be of clinical use possibly in confirm ing a p ren atal diagnosis m ade from karyotyping or in the case of the c e n tro m e ric 16 p ro b e to b e u s e d o n p r o d u c ts of c o n c e p tio n fo llo w in g m iscarriages. T risom y 16 is the m o st freq u en tly o b serv ed trisom y seen in sp o n tan eo u s abortions ( Kajii et al., 1986) and culturing of this m aterial has a very low success rate in this laboratory. FISH w ith probes for the m ost common
abnorm alities seen in abortions could provide som e insight as to the cause of som e of the m iscarriages from w hich sam ples are received in the diagnostic laboratory.
The centromeric 18 probe was used w ith lim ited success on seven blood smears follow ing fixing onto the slide. The hybridisation efficiency w as never m ore th an 33% on these sam ples. Since probe L I.84 has been used successfully on interphase cells from products of conception w ith 93 -100% of cells in m aterial from trisomie fetuses displaying three signals it w ould appear that the probe is efficient b u t cell preparation and hybridisation techniques were not optim al for the b lo o d sm ears. It w o u ld clearly be a d v an ta g e o u s to confirm a clinical d ia g n o s is of tris o m y 18 as w ell as to ex clu d e E d w a rd s sy n d ro m e fro m dysm orphic new born infants w ithin 24 hours of receiving a sample. N ew born infants w ith am biguous genitalia are also referred for k ary o ty p in g an d this situ atio n can be very distressing for the parents. A lth o u g h in u rg en t cases chrom osom es are prep ared from bloods w ithin 48 hours the m itotic index is frequently very low an d the quality of the m etaphases very poor. The area of utilising interphase FISH on blood smears w ithout intervening cell culture is one w hich should be investigated fu rth e r and it seem s likely that w ith im proved techniques it could become a valuable additional tool in the routine diagnostic laboratory.