Diagnosis of leukaemias – cytogenetic techniques
Following a short period of culturing the diagnostic sample, metaphase chromosomes are analysed to establish the karyotype. This assay requires a fresh heparinised bone marrow or blood sample.
Giemsa-banded metaphase after capture by an automated microscope reveals a classical t(9;22) translocation, as in CML (A).
Complex karyotypes are hard to decipher by standard banding, and 24-colour fluorescent in situ hybridisation (FISH) on the identical metaphase helps to resolve complex rearrangements (B).
In a routine workflow, cytomorphology and flow cytometry are rapid techniques that usually yield results within a few hours.
The typical morphology and immunophenotype can raise suspicion for certain subtypes of AML, which need further specification.
A conventional karyogram then returns the final diagnosis, e.g. an AML with a recurrent cytogenetic aberration: a t(8;21) translocation.
FISH is a tool to detect specific chromosomal aberrations. It can be applied to interphase nucleoli or metaphases after cell culture.
Probes are designed to bind specific genomic regions and allow the detection of trisomy (A), deletions (B) and translocations (C).
FISH is more sensitive than karyotyping and, in cases of specific translocations, can detect 1 in 200 cells (0.5%).
Revision Questions
- What is the purpose of conventional cytogenetics?
- What material (i.e. fresh or fixed) can be used for a cytogenetic workup?
- What is the lower sensitivity level of FISH?
