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Pitfalls in molecular diagnosis in haemato-oncology
  1. Joanne Mason,
  2. Susanna Akiki,
  3. Mike J Griffiths
  1. West Midlands Regional Genetics Laboratory, Birmingham Women's NHS Foundation Trust, Edgbaston, Birmingham, UK
  1. Correspondence to Joanne Mason, West Midlands Regional Genetics Laboratory, Birmingham Women's NHS Foundation Trust, Edgbaston, Birmingham B15 2TG, UK; joanne.mason{at}

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Molecular techniques can contribute to establishing the correct diagnosis, prognostic stratification and predicting and assessing response to treatment in haematological malignancy. For instance, presence of the JAK2V617F mutation confirms a suspected diagnosis of a myeloproliferative neoplasm (MPN), PML-RARA confirms acute promyelocytic leukaemia (APML), BCR-ABL1 confirms chronic myeloid leukaemia (CML) and FIP1L1-PDGFRA confirms chronic eosinophilic leukaemia (CEL).1 Examples of specific molecular markers on which targeted therapies depend include PML-RARA in APML, which responds to all-trans retinoic acid, and BCR-ABL1 in CML and FIP1L1-PDGFRA in chronic CEL which both respond to imatinib.

This brief review will illustrate with examples some of the common problems that may be encountered when using molecular techniques in the field of clinical molecular haemato-oncology.

Tissue type, sample quality and sensitivity issues

A major difference between genetic diagnosis of constitutional disorders and acquired neoplasia is the background of normal cells, which are invariably present in a tumour specimen. With blood cancer, this can be particularly problematic. For example, in systemic mastocytosis (SM), a MPN associated with a point mutation (D816V) in the KIT gene,2 the mutation may not be reliably detected in blood if there are insufficient mast cells.3 Literature discrepancies regarding the prevalence of KIT mutations in SM can be partly explained by differences among studies in the type of tissue screened. Marrow is recommended when referring samples for molecular diagnosis of SM,4 although cell selection using CD117 for enrichment of mast cells from blood may improve sensitivity.

The presence of normal cells is also an issue in the diagnosis of JAK2V617F-positive MPN, although the clinical significance of detecting a low level mutation-positive clone is questionable. Assays based on allele-specific oligonucleotide (ASO) PCR or amplification-refractory mutation system (ARMS) PCR have limited sensitivity, which can be compounded by the visualisation method (agarose gel electrophoresis being less sensitive than …

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  • Competing interests None.

  • Provenance and peer review Commissioned; externally peer reviewed.