Small marker chromosome identification in metaphase and interphase using centromeric multiplex FISH (CM-FISH)

Multicolor karyotyping procedures, such as multiplex fluorescence in situ h ybridization (M-FISH), spectral karyotyping, or color-changing karyotyping, can be used to detect chromosomal rearrangements and marker chromosomes in prenatal diagnosis. peripheral blood cultures, leukemia, acid solid tumors , especially in cases where G-banding is not sufficient. A regular M-FISH a nalysis requires relatively large amounts of labeled DNA (microgram quantit ies), is not informative in interphase nuclei, hybridization can take up to 2 to 3 days, and unlabeled human chromosome-painting probes are not availa ble commercially. Unique probes (plasmids, PAC), specific for centromeric o r subtelomeric chromosomal regions, can replace the painting probes in M-FI SH to address specific issues, such as the identification of marker chromos omes and aneuploidies. A set of plasmid probes carrying repetitive sequence s specific for the alpha -satellite region of all human chromosomes were co mbined in a metaphase assay and an interphase assay, allowing identificatio n of aneuploidies in one hybridization step, on a single cytogenetic slide. The fluorophore-dUTP and the labeled antibodies required to label and dete ct the DNA probes can be prepared in any laboratory. All DNA probes can be easily isolated and labeled using common molecular cytogenetic procedures. Because of the repetitive nature of the probes, hybridization time is short , usually less than 1 hour, and the analysis can be performed with nonspeci alized image-processing software.