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Chapter 21 : Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping

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Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, Page 1 of 2

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Abstract:

Acute lymphoblastic leukemia (ALL) and lymphoblastic lymphoma are malignant neoplasms of progenitor cells committed to B-lineage or T-lineage lymphopoiesis, termed lymphoblasts (1). Distinction between lymphoblastic leukemia (predominantly bone marrow with or without peripheral blood involvement) and lymphoblastic lymphoma (predominantly mass-forming nodal and/or extranodal disease) is largely clinical; however, the presence of greater than 25% bone marrow blasts generally prompts the designation of “leukemia” in most protocols, regardless of the presence of a mass lesion (2, 3). In the most recent World Health Organization (WHO) classification (2–4), B lymphoblastic leukemia/lymphoma is further subclassified on the basis of recurrent genetic abnormalities, while T lymphoblastic leukemia/lymphoma, despite the existence of recurrent genetic abnormalities, is not further subclassified (Table 1).

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Figures

Image of FIGURE 1
FIGURE 1

Immunophenotypic patterns of normal B-cell maturation in bone marrow. (A) Gated on all viable cells; B-cell maturational stages denoted by different colors. (B through D) Gated only on B cells; B-cell maturational stages denoted by different colors, from least mature (turquoise) to most mature (blue). (E and F) Acquisition of surface-membrane immunoglobulin light chain expression: (E) Gated on light chain-negative B cells (orange); (F) Gated on light chain-positive B cells: kappa (green), lambda (red).

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Image of FIGURE 2
FIGURE 2

Immunophenotypic patterns of normal T-cell maturation in thymus. (A through D). Gated on all cytoplasmic CD3 T cells; T-cell maturational stages denoted by different colors, from least mature (violet) to most mature (red and green). (E) Gated on sCD3- T cells. (F) Gated on sCD3 T cells.

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Image of FIGURE 3
FIGURE 3

Typical gating strategy in B lymphoblastic leukemia. (A) All events displayed; a gate defining single events is illustrated. (B) All single events resulting from the gate shown in panel A are displayed; a gate defining viable cells is illustrated. (C) All viable single events are displayed; a gate defining all CD19 B cells is illustrated. (D) All viable, single CD19 B cells are displayed; blasts (blue) may be distinguished from mature B cells (yellow) by gating on CD45-negative to CD45 events.

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Image of FIGURE 4
FIGURE 4

Example of B lymphoblastic leukemia. Phenotypic patterns of neoplastic B lymphoblasts (blue, gated as in Fig. 3 ) are compared with those of normal bone marrow B cells (pink).

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Image of FIGURE 5
FIGURE 5

Example of T lymphoblastic leukemia. Phenotypic patterns of neoplastic T lymphoblasts (yellow) are compared with those of normal thymic T cells (blue).

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Image of FIGURE 6
FIGURE 6

Example of early T-cell precursor (ETP) leukemia. In this example, the neoplastic T lymphoblasts (violet) are positive for surface-membrane CD3, with weak CD5 expression in significantly less than 75% of the blasts. CD1a and CD8 are negative, and at least 25% of the blasts express one or more myeloid or stem-cell marker (in this case, CD34). Normal thymic T cells are shown for comparison in blue.

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Image of FIGURE 7
FIGURE 7

Detection of minimal residual disease in B lymphoblastic leukemia. (A) All viable, single CD19 bone marrow B cells are displayed; a small cluster of cells (0.04% of viable singlets) with comparatively dim CD45 expression (green) can be distinguished from a predominant population of normal B cells (blue). (B through F) Phenotypic features of the minimal residual disease (green, gated as in panel A) are compared with those of normal bone marrow B cells (pink). Compared with normal B-cell precursors, the residual neoplastic cells express abnormally bright CD10, CD19, CD49f, CD58, CD86, and CD304 and are further positive for CD13 and/or CD33. CD38 expression, moreover, is lost.

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Image of FIGURE 8
FIGURE 8

Detection of minimal residual disease in T lymphoblastic leukemia. (A) All viable, single bone marrow T cells are displayed; a small cluster of cells (0.2% of viable singlets) with loss of surface CD3 (violet) can be distinguished from a predominant population of normal sCD3 T cells (yellow). (B through D) The phenotypic features of the minimal residual disease are compared with those of normal thymic T cells (light blue) and the normal mature (sCD3) T cells contained in the bone marrow sample (yellow). With respect to absence of sCD3 (B) and weak coexpression of CD4 and CD8 (C), the neoplastic T cells resemble normal thymic T cells; however, T cells with a thymic immunophenotype are not detectable in normal bone marrow by flow cytometry. The combination of weak CD5 and intermediate CD45 seen in the minimal residual disease (D), though, differs from both normal thymic T cells and mature T cells. The residual leukemic cells were also partially positive for CD1a and CD10 (not shown).

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
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Tables

Generic image for table
TABLE 1

World Health Organization classification of precursor lymphoid neoplasms (2 )

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
Generic image for table
TABLE 2

Sample antibody combinations and applications

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21
Generic image for table
TABLE 3

Immunophenotypic-genotypic associations

Citation: DiGiuseppe J. 2016. Acute Lymphoblastic Leukemia/Lymphoma: Diagnosis and Minimal Residual Disease Detection by Flow Cytometric Immunophenotyping, p 207-216. In Detrick B, Schmitz J, Hamilton R (ed), Manual of Molecular and Clinical Laboratory Immunology, Eighth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818722.ch21

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