PURPOSE: Treatment stratification in ALL includes diverse (cyto)genetic aberrations, requiring diverse tests to yield conclusive data. We optimized the diagnostic workflow to detect all relevant aberrations with a limited number of tests in a clinically relevant time frame. METHODS: In 467 consecutive patients with ALL (0-20 years), we compared RNA sequencing (RNAseq), fluorescence in situ hybridization (FISH), reverse transcriptase polymerase chain reaction (RT-PCR), karyotyping, single-nucleotide polymorphism (SNP) array, and multiplex ligation-dependent probe amplification (MLPA) for technical success, concordance of results, and turnaround time. RESULTS: To detect stratifying fusions ( CONCLUSION: Combining RNAseq and SNP array outperformed current diagnostic tools to detect all stratifying genetic aberrations in ALL. The turnaround time is <
15 days matching major treatment decision time points. Moreover, combining RNAseq and SNP array has the advantage of detecting new lesions for studies on prognosis and pathobiology.