This paper is a review of current diagnostic applications of molecular methods in pediatric hematooncology, including analyses performed at disease presentation, evaluation of prognosis as well as those for assessment of treatment effectiveness. Here we present the examples of important fields of application of molecular methods in pediatric hematooncology, i.e. identification of clinically significant fusion genes in acute lymphoblastic leukemia and monitoring of minimal residual disease in this most frequent childhood malignancy. Moreover, we present the methodology and clinical significance of quantitative analysis of hematopoietic chimerism after allogeneic stem cell transplantation. The methods presented include fluorescent in situ hybridization, reverse transcription, conventional and quantitative polymerase chain reaction, as well as methods of genotyping based on analysis of microsatellite marker polymorphism, single nucleotide polymorphism, and Indel markers.