After approximately 20 years of development and after several prospective clinical trials, the detection of minimal residual disease (MRD) has emerged as part of state-of-the-art diagnostics to guide the majority of contemporary treatment programs both in pediatric and adult acute lymphoblastic leukemia (ALL). For ALL, several methods of MRD analysis are available, but 2 are widely applicable. One is based on the detection of aberrant expression of leukemia specific antigens by flow cytometry and the other one uses the specific rearrangements of the TCR or Ig genes, which can be detected by quantitative PCR in the DNA of leukemic cells. In some cases with known fusion genes such as BCR/ABL, RT-PCR can be used as a third method of identifying leukemic cells by analyzing RNA in patient samples. Clinical application of such sophisticated tools in the stratification and treatment of ALL requires reliable, reproducible, and quality-assured methods to ensure patient safety.