Corrigendum: Dogs with acute myeloid leukemia have clonal rearrangements in T and B cell receptors

The tests used to define criteria were run sequentially (Figure 1), but the combined results were evaluated after completion of analysis to obtain a definitive diagnosis of AML.

ALP, alkaline phosphatase; AML, acute myeloid leukemia; ANBE, α-naphthyl butyrate esterase; CAE, chloroacetate esterase; MPx, myeloperoxidase; SBB, Sudan Black B.

In some dogs, the final classification was presumptive (suspect) because bone marrow was not available for the classification assays (flow cytometric analysis and cytochemical staining) or one of the classification assays was not done.

^aBecause both neutrophils and monocytes can express CD4 (with negative T cell markers), CD11b, or CD11c, these markers support a diagnosis of AML but alone do not differentiate between subtypes (other criteria were used instead) and are not listed here.

AF, alexafluor; PC, allophycocyanin; FITC, fluorescein isothiocyanate; PE, phycoerythrin; pmn, neutrophil.

^aAbcam, Cambridge, MA, USA; AbD Serotec, now part of Bio-Rad, Hercules, CA, USA; BD-Biosciences, Franklin Lakes, NJ, USA; Beckman-Coulter, Fullerton, CA, USA; Dako, now part of Agilent Technologies, Santa Clara, CA, USA; UC-Davis: Peter Moore, University of California-Davis, Leukocyte Antigen Biology Laboratory, Davis, CA, USA; VMRD, Pullman, WA, USA.

All results were obtained from testing done at Cornell University unless stated otherwise in parentheses. Note that not all of the antibodies provided in Table 3 were used in every dog, and negative results for flow cytometric analysis was confined to T (CD3, CD5, TCRαβ), B (CD21, CD22), or stem (CD34) cell markers, the pan-leukocyte marker (CD18), and MHCII. If results for the latter markers are not provided, then the antibody against the marker was not tested in that case. If a marker or cytochemical reaction is not listed, it means the test result was negative (usual scenario) or not performed. When available, the percentage of blasts with positive cytochemical reactions (and the strength of the reaction for ALP) is provided. NA, not available for review with no report provided; ND, not done; PLF, pleural fluid; PTF, peritoneal fluid; M4, acute myeloid leukemia – not otherwise specified, myelomonocytic; M5, acute myeloid leukemia – not otherwise specified, monocytic or monoblastic; CSU, Colorado State University; pmn, neutrophil; mono, monocyte; eos, eosinophil; IHC, immunohistochemical; VB, venous blood; AML, acute myeloid leukemia; BM, bone marrow; ALP, alkaline phosphatase; ANBE, α-naphthyl butyrate esterase; CAE, chloroacetate esterase; MPx, myeloperoxidase. Underlined dog numbers were included in our previous publication on ALP staining in AML (16). Case 2 was also presented in the February 2015 American Society of Veterinary Clinical Pathology on-line rounds (available to members only at https://www.asvcp.org/).

^aNo positive reactions were observed for MPx in those cases in which this cytochemical staining.

^bAdditional rationale for classification of AML subtype is provided: (1) for mixed lineage #2, flow cytometric and cytochemical staining results in bone marrow supported a monoblastic lineage, but negative results for cytochemical stains and morphologic features supported an erythroid lineage in spleen; (2) for suspect M5 #4, the cells lacked flow cytometric markers of myeloid differentiation and only expressed strong ALP on cytochemical staining, which is not expressed in normal mature or immature neutrophils in dogs and supports an AML (16). The lack of expression of the neutrophil enzyme CAE in blasts on cytochemical staining argues against a myelomonocytic classification despite the presence of ≥20% mature and immature neutrophils in blood (bone marrow not available for evaluation); (3) for suspect M5 #9, morphologic features and strong positive ALP staining supported monocytic differentiation; (4) for suspect M4 #11, the cells expressed neutrophil or monocyte differentiation antigens on flow cytometry and the percentage of CAE- positive cells on cytochemical staining was close to 20%; (5) for mixed lineage #20, there was concurrent expression of B (CD22, this was not confirmed by IHC for Pax-5 on a core biopsy) and T (CD3, this was not confirmed on IHC staining of a core biopsy with CD3) cell markers with flow cytometry and myeloid markers on cytochemical staining (diffuse light or chunky ANBE, CAE positive); (6) for mixed lineage #23, there was morphologic (micromegakaryocytes), flow cytometric (CD61), and cytochemical evidence (diffuse light or chunky ANBE) of megakaryocytic differentiation, but cells also expressed ALP (lacking in normal megakaryocytes and platelets in the dog) and showed morphologic features of myeloid differentiation. The tumor cells also expressed CD3.

^c#21: limited flow cytometric panel done (only conjugated antibodies); #22: insufficient blasts on the blood smear to evaluate 100 cells on cytochemical staining; #25: flow cytometric analysis on the lymph node was done at CSU at presentation to the oncologist. The dog was euthanized 2.5 months after diagnosis of and treatment for AML and venous blood and bone marrow were submitted to Cornell University for morphologic assessment, flow cytometric analysis, and cytochemical staining. At the time of euthanasia, the dog had a total leukocyte count of 79.8 × 10⁶/L, consisting of 39.1 × 10⁶/L blasts (a few with purple cytoplasmic granules), with 32.7 × 10⁶/L monocytes (some dysplastic) and 0.8 × 10⁶/L neutrophils in blood.