Juvenile myelomonocytic leukemia presenting in an infant with a subdural hematoma.

Background: Juvenile myelomonocytic leukemia (JMML) is a rare childhood hematopoietic disorder typically presenting with hepatosplenomegaly, lymphadenopathy, pallor, fever, and cutaneous findings. The authors report the first case, to our knowledge, of JMML presenting in a pediatric patient with a subdural hematoma.

Case Description: A 7-month old male with recurrent respiratory infections and a low-grade fever presented with a full fontanelle and an increasing head circumference and was found to have chronic bilateral subdural collections.

CD49f protein expression varies among genetic subgroups of B lymphoblastic leukemia and is distinctly low in KMT2A-rearranged cases.

Background: CD49f (integrin α6) is a useful marker for minimal residual disease (MRD) detection in B lymphoblastic leukemia and has recently been suggested to mediate infiltration of the central nervous system by leukemic B lymphoblasts. However, data regarding expression of CD49f protein in B lymphoblastic leukemia are limited, and whether CD49f protein expression varies among genetic subgroups of B lymphoblastic leukemia is unknown.

Methods: CD49f protein expression was characterized by flow cytometry in a series of 40 cases of B lymphoblastic leukemia, which included the genetic subgroups: KMT2A-rerranged, BCR-ABL1+, ETV6-RUNX1+, hypodiploidy, and hyperdiploidy.

Immunophenotyping of Acute Lymphoblastic Leukemia.

Immunophenotyping by flow cytometry is an important component in the diagnostic evaluation of patients with acute lymphoblastic leukemia. This technique further permits the detection of minimal residual disease after therapy, a robust prognostic factor that may guide individualized treatment. We describe here laboratory methods for both the initial characterization of lymphoblasts at diagnosis, and the detection of rare leukemic lymphoblasts after treatment.

Intracytoplasmic crystalline inclusions in chronic lymphocytic leukemia.

Neoplastic B cells may have cytoplasmic inclusions that are visible in routine peripheral blood smears by light microscopy.

Applications of Flow Cytometric Immunophenotyping in the Diagnosis and Posttreatment Monitoring of B and T Lymphoblastic Leukemia/Lymphoma.

In this review, we discuss applications of flow cytometric immunophenotyping (FCI) in the diagnostic evaluation and posttreatment monitoring of B and T lymphoblastic leukemia/lymphoma. We describe practical approaches to FCI at the time of diagnosis, with an emphasis on blast identification, lineage assignment, and distinction of B and T lymphoblastic leukemia/lymphoma from their morphologic and immunophenotypic mimics. We further review flow cytometric assays for the detection of minimal or measurable residual disease (MRD) after treatment, and illustrate both standard approaches, and newer strategies for improving sensitivity and circumventing the loss of immunophenotypic targets after immunotherapy.

Breast implant-associated anaplastic large-cell lymphoma.

In patients with suspected breast implant-associated anaplastic large-cell lymphoma, cytologic evaluation of fine-needle aspirate specimens from the peri-implant seroma, together with flow cytometric immunophenotyping and immunohistochemistry, represents a suitable preoperative diagnostic approach when planning for surgical management.

PhenoGraph and viSNE facilitate the identification of abnormal T-cell populations in routine clinical flow cytometric data.

Background: Flow cytometric identification of neoplastic T-cell populations is complicated by the wide range of phenotypic abnormalities in T-cell neoplasia, and the diverse repertoire of reactive T-cell phenotypes. We evaluated whether a recently described clustering algorithm, PhenoGraph, and dimensionality-reduction algorithm, viSNE, might facilitate the identification of abnormal T-cell populations in routine clinical flow cytometric data.

Methods: We applied PhenoGraph and viSNE to peripheral blood mononuclear cells labeled with a single 8-color T/NK-cell antibody combination.

A QA Program for MRD Testing Demonstrates That Systematic Education Can Reduce Discordance Among Experienced Interpreters.

Background: Minimal residual disease (MRD) in B lymphoblastic leukemia (B-ALL) by flow cytometry is an established prognostic factor used to adjust treatment in most pediatric therapeutic protocols. MRD in B-ALL has been standardized by the Children's Oncology Group (COG) in North America, but not routine clinical labs. The Foundation for National Institutes of Health sought to harmonize MRD measurement among COG, oncology groups, academic, community and government, laboratories.

Risk stratification of chromosomal abnormalities in chronic myelogenous leukemia in the era of tyrosine kinase inhibitor therapy.

Clonal cytogenetic evolution with additional chromosomal abnormalities (ACAs) in chronic myelogenous leukemia (CML) is generally associated with decreased response to tyrosine kinase inhibitor (TKI) therapy and adverse survival. Although ACAs are considered as a sign of disease progression and have been used as one of the criteria for accelerated phase, the differential prognostic impact of individual ACAs in CML is unknown, and a classification system to reflect such prognostic impact is lacking. In this study, we aimed to address these questions using a large cohort of CML patients treated in the era of TKIs.

Detection of minimal residual disease in B lymphoblastic leukemia using viSNE.

Background: Minimal residual disease (MRD) following treatment is a robust prognostic marker in B lymphoblastic leukemia. However, the detection of MRD by flow cytometric immunophenotyping is technically challenging, and an automated method to detect MRD is therefore desirable. viSNE, a recently developed computational tool based on the t-Distributed Stochastic Neighbor Embedding (t-SNE) algorithm, has been shown to be capable of detecting synthetic "MRD-like" populations of leukemic cells created in vitro, but whether viSNE can facilitate the immunophenotypic detection of MRD in clinical samples has not been evaluated.

Detection of small abnormal B-Lymphoblast populations at diagnosis of chronic myelogenous leukemia, BCR-ABL1+: Incidence, phenotypic features, and clinical implications.

Background: According to the 2008 World Health Organization (WHO) Classification of Tumors of the Haematopoietic and Lymphoid Tissues, the finding of B lymphoblasts in the blood or bone marrow of a patient with chronic myelogenous leukemia, BCR-ABL1+ (CML) should raise a concern for progression of the disease to B-lymphoblastic blast phase. Data addressing the incidence and phenotypic features of abnormal B lymphoblasts in CML, and whether the detection of B lymphoblasts inexorably heralds blast phase in CML, though, are limited.

Methods: We reviewed a consecutive series of patients with newly diagnosed CML who had undergone bone marrow examination with flow cytometric immunophenotyping.

Detection of B-cell populations with monotypic light chain expression in cerebrospinal fluid specimens from patients with multiple sclerosis by polychromatic flow cytometry.

Background: Flow cytometric immunophenotyping is an established method for the detection of occult leptomeningeal disease in patients with aggressive B-cell non-Hodgkin lymphoma, and is increasingly being used in the evaluation of patients without an established diagnosis of lymphoma who present with signs and/or symptoms referable to the central nervous system. However, the specificity of flow cytometric immunophenotyping in the identification of monoclonal B-cell populations in cerebrospinal fluid (CSF) has not been systematically evaluated.

Methods: We searched a consecutive series of CSF specimens submitted to our laboratory for polychromatic (8-color) flow cytometric immunophenotyping between June, 2010 and December, 2012 for cases in which a B-cell population with monotypic immunoglobulin light chain expression was detected in patients without clinical or radiographic evidence of lymphoma.

Improved compensation of the fluorochrome AmCyan using cellular controls.

Background: Implementation of polychromatic flow cytometry in the clinical laboratory often requires the use of newer fluorochromes, with which experience may be comparatively limited. In the course of implementing polychromatic flow cytometry in our laboratory, we have observed significant differences in compensation values derived for the violet-excited dye, AmCyan, when cells rather than a commercially available set of polystyrene microparticles (BD CompBeads) are used as compensation controls.

Methods: Compensation values were calculated for AmCyan and several other fluorochromes using the BD CompBeads Set according to the manufacturer's protocol, and using unstained and singly stained lymphocytes as compensation controls.

Guidelines for the diagnosis and monitoring of paroxysmal nocturnal hemoglobinuria and related disorders by flow cytometry.

Background: Paroxysmal nocturnal hemoglobinuria (PNH) is a rare hematopoietic stem cell disorder characterized by a somatic mutation in the PIGA gene, leading to a deficiency of proteins linked to the cell membrane via glycophosphatidylinositol (GPI) anchors. While flow cytometry is the method of choice for identifying cells deficient in GPI-linked proteins and is, therefore, necessary for the diagnosis of PNH, to date there has not been an attempt to standardize the methodology used to identify these cells.

Methods: In this document, we present a consensus effort that describes flow cytometric procedures for detecting PNH cells.

Overexpression of CD49f in precursor B-cell acute lymphoblastic leukemia: potential usefulness in minimal residual disease detection.

Background: The persistence of minimal residual disease (MRD) following therapy is an established prognostic factor in precursor B-cell acute lymphoblastic leukemia (pB-ALL). Detection of MRD in pB-ALL by flow cytometric immunophenotyping requires demonstration of abnormal antigen expression in leukemic B-cell precursors relative to that of normal B-cell precursors. The gene encoding CD49f (integrin alpha-6) is one of several whose overexpression in pB-ALL at diagnosis has been associated with the subsequent detection of MRD.

Racial/ethnic variation in EBV-positive classical Hodgkin lymphoma in California populations.

Epstein-Barr virus (EBV) is detected in the tumor cells of some but not all Hodgkin lymphoma (HL) patients, and evidence indicates that EBV-positive and -negative HL are distinct entities. Racial/ethnic variation in EBV-positive HL in international comparisons suggests etiologic roles for environmental and genetic factors, but these studies used clinical series and evaluated EBV presence by differing protocols. Therefore, we evaluated EBV presence in the tumors of a large (n = 1,032), racially and sociodemographically diverse series of California incident classical HL cases with uniform pathology re-review and EBV detection methods.