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Chromosome band specific FISH probes allow improved detection of terminal translocations in leukaemic metaphases

Leukemia volume 15, pages 860–861 (2001)Cite this article

Fluorescent in situ hybridisation (FISH) probes capable of detecting the entire length of individual chromosomes (whole chromosome paints) have become a valuable and widely used tool for identifying large structural and numerical abnormalities, as well as cryptic abnormalities beyond the resolution of G-banding.1 It has previously been established, however, that whole chromosome painting probes (WCPs) have a limited resolution, and fail to detect small chromosomal segments, such as those involved in subtle translocations with breakpoints located in the terminal band.2 One such rearrangement is the Philadelphia translocation t(9;22)(q34.1;q11.2) in chronic myeloid leukaemia (CML), in which the majority of chromosome 22q is transferred to chromosome 9 while a relatively small amount of chromosome 9 material relocates to chromosome 22. While chromosome 22 material is clearly visible on the derivative chromosome 9 [der(9)] by both G-banding and whole chromosome painting, the small fragment of chromosome 9 present on the der(22), or Philadelphia (Ph) chromosome, is beyond the resolution of G-banding and difficult to detect even with the application of whole chromosome paints. Our experience has shown that WCP 9 fails to detect chromosome 9 material on the Ph chromosome in up to 90% of cells. In metaphases where a WCP 9 signal is visible on the Ph chromosome the hybridisation signals are small and faint. This apparent lack of sensitivity of WCP 9 is observable in previously published FISH images of Ph positive metaphases.3 Recently the versatility of chromosome painting has improved with the development of the band specific painting probe (BSP). BSPs are amplified from microdissected chromosome material and fluorescently labelled to allow detection of these subchromosomal regions. We have found that BSPs provide greater sensitivity than WCPs even when applied to late metaphase figures. For example, in CML patients a band specific probe for region 9q34 provides a strong and reliable signal on the Philadelphia chromosome. To illustrate the sensitivity of band specific probes we present our results on metaphases from a CML patient carrying a variant Ph translocation. The location of band 9q34 in this patient could only be elucidated after application of 9q34 BSP.

Cytogenetic analysis was performed on metaphase chromosomes from unstimulated bone marrow short-term cultures, G-banded with a conventional trypsin-Giemsa technique. BCR-ABL detection was carried out using LSI BCR/ABL (Vysis Inc, Downers Grove, IL, USA). WCPs 5, 9 (Cambio, Cambridge, UK) and 22 (Vysis) were applied according to the manufacturer's instructions. BSP 9q34 (Research Genetics, Inc, Huntsville, AL, USA) was amplified using a degenerate oligonucleotide primed-PCR method,4 and labelled by the same method using Cy3-conjugated primers (MWG Biotech, Bedfordshire, UK).

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References

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Acknowledgements

This work was supported by the Leukaemia Research Fund and the Kay Kendal Leukaemia Fund.

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  1. Department of Haematology, University of Cambridge,

    A Reid, SM Gribble, AR Green & EP Nacheva

  2. Department of Haematology, Addenbrook's Hospital, Cambridge, UK

    KM Andrews

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  1. A Reid
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  2. SM Gribble
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Reid, A., Gribble, S., Andrews, K. et al. Chromosome band specific FISH probes allow improved detection of terminal translocations in leukaemic metaphases. Leukemia 15, 860–861 (2001). https://doi.org/10.1038/sj.leu.2402096

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