Accurate immunophenotyping and cytogenetic analysis of haematological malignancies or blood cancers are essential at diagnosis for deriving appropriate therapeutic strategies. Bone marrow specimens are obtained to detect the presence of an abnormal number, distribution or type of cells. This involves immunophenotyping the bone marrow for specific neoplastic cell populations and manual estimation of the number of antigen-positive cells by microscopy, which is highly inaccurate. Assessment of phenotyped cells by flow cytometry is more accurate but lacks information on cellular location within the marrow and is unable to detect genetic abnormalities. Cytogenetic analysis by fluorescence in situ hybridisation (FISH) informs prognosis and is an integral component when determining patient treatment. Currently, FISH is performed manually on cells from bone marrow and blood biopsies. This is labour intensive and has low sensitivity with the nuclei of only 100-200 cells analysed. This is not suitable when the number of neoplastic cells is low, such as for monitoring residual disease following treatment.
The Translational Cancer Pathology Laboratory at the University of Western Australia is applying new imaging techniques to increase the sensitivity of blood cancer diagnosis. Automated digital enumeration of immunohistochemically stained cells was applied to microscopy assessment and found to increase sensitivity. We have further improved sensitivity by implementing image-based flow cytometry which combines high resolution digital images with the quantitative fluorescence information gained from a standard flow cytometer. Combining immunophenotyping with FISH in a single test enables genetic abnormalities to be directly assessed in neoplastic cells identified by phenotype. This provides accurate analysis of chromosomal abnormalities such as changes in copy number (aneuploidy) or locus deletion (eg TP53). This demonstrates that imaging flow cytometry has the capacity to bridge the gap that currently exists between the diagnostic tests that detect important phenotypic and genetic changes in the clinical assessment of blood cancers.