Development of a probe consist of three cosmids to enumerate the chromosome 13 on uncultured lymphocytes or amniocytes using interphase FISH

 Abstract Background: To produce a reliable probe suitable for aneuploidy detection of chromosome 13 on uncultured lymphocytes and amniocytes by fluorescence in situ hybridization (FISH), we used a contig of three overlapping cosmids mapped to 13q12.3. Methods: The cosmid DNA carrying the expected sequences of human chromosome 13 was isolated from host cells and labelled with biotin-11-dUTP. The hybridization and detection conditions with FITC-Avidin were optimised using a series of cultured and uncultured lymphocytes and amniocytes. Results: Intensive signals were detected when a combination of three overlapping cosmids was used to enumerate the chromosome 13 on interphase nuclei. An average of 87 and 85.5 percent of interphase cells prepared from lymphocytes and amniocytes showed accurate number of specific signals for chromosome 13. Conclusion: The results obtained in present study indicate that the probe was capable of detecting the copy number of chromosome 13 on interphase cells prepared from peripheral blood or amniotic fluid cells providing that the uncultured amniotic fluid cells are free of cytoplasmic residues, RNA and protein debris.