Acute Lymphoblastic Leukemia - ALL

Last Literature Review: September 2022 Last Update:

Medical Experts


Acute lymphoblastic leukemia (ALL) is an aggressive type of leukemia of either B- or T-lineage immature lymphoid cells. The prognosis for patients with ALL can be inferred from age and white blood cell (WBC) count at diagnosis. An initial ALL workup generally involves a CBC with peripheral smear, flow cytometric immunophenotyping, coagulation studies, liver function testing, and chemistry profiles. A definitive diagnosis of ALL often requires morphologic demonstration of increased lymphoblasts within bone marrow aspirate or biopsy samples.   Cytogenetic studies and fluorescence in situ hybridization (FISH) analysis allow for the identification of genetic abnormalities that inform risk stratification, treatment planning, and monitoring.  

Quick Answers for Clinicians

Which tests are used to diagnose acute lymphoblastic leukemia?

The minimum diagnostic workup for acute lymphoblastic leukemia (ALL) includes a morphologic examination of peripheral blood (and almost always bone marrow), immunophenotyping via flow cytometry, cytogenetics, and molecular characterization of leukemic cells (eg, via fluorescence in situ hybridization [FISH] or molecular genetic testing).  These studies allow for the diagnosis and characterization of the ALL subtype based on combined cytogenetic and immunophenotypic characteristics of lymphoblasts. The results of these tests also determine appropriate treatment protocols and provide a basis for subsequent detection of minimal residual disease (MRD).    

What role does next generation sequencing play in acute lymphoblastic leukemia testing?

Clonal rearrangements of the B- and/or T-cell receptor genes may be used to aid in the detection of minimal/measurable residual disease (MRD). 

What pediatric-specific acute lymphoblastic leukemia guidance is available?

The National Comprehensive Cancer Network (NCCN) provides guidance concerning acute lymphoblastic leukemia (ALL) in pediatric patients. The NCCN recommendations are categorized by risk level, which can be age related, and discuss long-term supportive care. Refer to the NCCN’s Pediatric Acute Lymphoblastic Leukemia guidelines for more information. 

Indications for Testing

ALL testing should be considered in patients who present with signs of bone marrow failure (eg, anemia, thrombocytopenia, leukopenia) and constitutional symptoms (eg, fever, lethargy, weight loss).  In children, joint or extremity pain may be the only presenting symptom.  

Laboratory Testing

Initial Workup

The recommended laboratory testing in the initial workup of suspected ALL includes a CBC with a platelet count and automated differential, peripheral blood smear, chemistry profiles, infectious disease testing, liver function tests, tests for disseminated intravascular coagulation (DIC), tests for tumor lysis syndrome, infectious disease testing, and pregnancy testing.  


Required testing for the diagnosis of ALL includes morphologic evaluation of bone marrow to determine blast counts, flow cytometric immunophenotyping to define the type of leukemia, and molecular characterization of leukemic cells (eg, cytogenetic evaluation, FISH analysis, and/or molecular genetic testing) to define risk-associated biologic drivers of leukemia.  In clinical scenarios that prohibit bone marrow examination, these studies may be performed on peripheral blood samples, provided there is a sufficient blast count.  Cumulatively, these studies enable characterization, prognostication, and treatment planning.     In patients with diagnosed ALL, it is also important to obtain a cerebrospinal fluid (CSF) sample; ALL may involve the central nervous system (CNS) both at diagnosis and at relapse. Knowledge of CSF status at diagnosis directs appropriate therapy. 


Flow cytometric immunophenotyping on bone marrow aspirate material is performed to identify certain cell surface markers that define leukemia subtypes and to predict patient outcomes.   If the bone marrow aspirate cannot be obtained or used, a core biopsy or peripheral blood (if there are enough blasts) may be tested. 

ALL is broadly classified into two immunophenotypic groups, B-cell ALL and T-cell ALL, based on flow cytometric immunophenotyping. Rare acute leukemias (acute leukemias of ambiguous lineage) may express an ambiguous phenotype, which precludes definitive lineage assignment. 

Cytogenetics and Molecular Testing

Cytogenetic and molecular tests are an important part of the diagnosis, prognosis, and treatment of ALL in pediatric and adult patients.  These tests further characterize ALL subtypes and provide prognostic stratification, especially in children.

Testing methods include karyotyping, FISH, and molecular tests such as single nucleotide polymorphism (SNP) microarray and next-generation sequencing (NGS, also referred to as massively parallel sequencing).

Karyotyping, or chromosome analysis, is used to identify recurrent cytogenetic abnormalities and is recommended in all cases of diagnosed or suspected ALL.   Karyotyping detects important abnormalities such as aneuploidy or chromosomal rearrangements.  

FISH is more sensitive than karyotyping in detecting cytogenetic abnormalities and is recommended (with appropriate probes) for the detection of the most common cytogenetic abnormalities.  Abnormalities that cannot be detected by karyotyping, but can be detected by FISH, include ETV6-RUNX1 and iAMP21. 

Microarray testing can detect some abnormalities that may be missed by chromosome analysis or FISH, such as focal deletions, but it will not detect balanced translocations. Microarray tests may be considered in addition to conventional cytogenetics, especially if karyotyping is not successful or if there is aneuploidy. 

In B-ALL, the National Comprehensive Cancer Network (NCCN) recommends testing for all relevant fusion genes and pathogenic variants via NGS to determine prognosis and plan treatment.  Either bone marrow or peripheral blood (with sufficient lymphoblasts) may be used. 

The BCR-ABL1 fusion gene [t(9;22)(q34.1;q11.2), also referred to as BCR::ABL1, the Philadelphia chromosome, or Ph] is an especially important abnormality in B-ALL. In addition to karyotyping, qualitative or quantitative reverse transcription polymerase chain reaction (RT-PCR) testing that includes transcript size (p190 or p210) determination is recommended. 

Monitoring During Treatment

Therapeutic response monitoring is performed to assess treatment efficacy and determine risk stratification. Minimal residual disease (MRD) evaluation is performed for nearly all pediatric and many adult patients with ALL. MRD assessment may include flow cytometry, quantitative RT-PCR (such as for the BCR-ABL1 p190 or p210 transcript), or NGS-based evaluation for B-cell and/or T-cell clonality. 


Thiopurine S-methyltransferase (TPMT) activity testing, TPMT genetic testing, and NUDT15 genetic testing are available to guide appropriate thiopurine dosing and mitigate the risk of toxicity.   Refer to the Clinical Pharmacogenetics Implementation Consortium (CPIC) Guideline for Thiopurines and TPMT and NUDT15. 

ARUP Laboratory Tests


Flow Cytometry

Specimens: bone marrow, whole blood, tissue, fluid



Specimen: bone marrow

Specimen: bone marrow

Specimen: whole blood

Specimen: whole blood


Specimens: bone marrow, whole blood

Probes detect BCR-ABL1 t(9;22), KMT2A (MLL) 11q23 rearrangement (partner not determined), TCF3 (E2A) rearrangement (partner not determined), IGH rearrangement (partner not determined), and MYC rearrangement

Specimens: bone marrow, whole blood

Probes detect BCR-ABL1 t(9;22), KMT2A (MLL) 11q23 rearrangement (partner not determined), ETV6-RUNX1, t(12;21), CEP4, and CEP10

Specimens: bone marrow, whole blood

Probes included: CRLF2, JAK2, EPOR, CSF1R, ABL1, ABL2, and PDGFRB

Specimens: bone marrow, whole blood

SNP Microarray

Specimens: bone marrow, peripheral blood

Molecular Testing for BCR-ABL1 (Ph+) Leukemia


Specimens: bone marrow, whole blood

Specimens: bone marrow, whole blood

Specimens: bone marrow, whole blood