Cytogenetics plays an important role in the diagnosis of acute lymphoblastic leukaemia (ALL), particularly in relation to the association of specific chromosomal abnormalities with outcome. The karyotype at diagnosis is used in the risk stratification of patients for treatment within trial-based protocols. Chromosomal analysis of the leukaemic cells of patients with ALL is challenging as the mitotic index may be low and the chromosomal morphology is often poor. Therefore, the use of fluorescence in situ hybridisation (FISH) in parallel with cytogenetic analysis is important for the detection of those chromosomal abnormalities of prognostic significance. This chapter is dedicated to the preparation of ALL samples for cytogenetic and FISH analysis, with emphasis on the modifications to standard protocols which may be used to improve their quality. The specific difficulties encountered in the analysis of ALL metaphases and suggestions for overcoming them are provided. The chapter also includes an overview of the abnormalities that are expected to be found in this disease and how the results from both cytogenetics and FISH should be interpreted.