-deletions occur recurrently in acute lymphoblastic leukemia, especially in the -rearranged subtype. The -deletion was shown to positively impact prognosis of patients with -deletion and its presence precludes assignment into group, a novel high-risk category on AIEOP-BFM ALL trials. We analyzed the impact of different methods on -deletion detection rate, evaluated -deletion as a potential marker for -rearranged leukemia, studied its associations with molecular and clinical characteristics within this leukemia subtype, and analyzed its clonality. Using single-nucleotide-polymorphism array, genomic polymerase chain reaction (PCR) and amplicon-sequencing we found -deletion in 34% (16 of 47), 66% (33 of 50) and 78% (39 of 50) of -rearranged leukemia, respectively. False negativity of -deletion by single-nucleotide-polymorphism array caused misclassification in 5 patients. No -deletion was found outside the -rearranged cases. Within -rearranged leukemia, the -deletion was associated with higher total number of copy-number aberrations, and, importantly, the -deletion positivity by PCR was associated with better outcome 5-year event-free survival (EFS), -deletion-positive 93% -deletion-negative 68%, =0.022; 5-year overall survival (OS), -deletion-positive 97% -deletion-negative 75%, =0.029. Ultra-deep amplicon-sequencing revealed distinct co-existing -deletions in 22 of 24 patients. In conclusion, our data demonstrate inadequate sensitivity of single-nucleotide-polymorphism array for -deletion detection, unacceptable for proper classification. Even using more sensitive methods (PCR/amplicon-sequencing) for its detection, -deletion is absent in 22-34% of -rearranged leukemia and does not represent an adequately sensitive marker of this leukemia subtype. Importantly, the -deletion potentially stratifies the -rearranged leukemia into biologically/clinically distinct subsets. Frequent polyclonal pattern of -deletions shows that late origin of this lesion is more common than has been previously described.