Int J Hematol DOI 10.1007/s12185-014-1558-2
IMAGES IN HEMATOLOGY
Ethylenediaminetetraacetate-dependent pseudothrombocytopenia accompanied by morphological changes Weiyi Xu • Baode Chen • Yu Chen
Received: 25 December 2013 / Revised: 20 February 2014 / Accepted: 20 February 2014 Ó The Japanese Society of Hematology 2014
A 65-year-old female patient was referred to our hospital for diabetes mellitus, diabetic nephropathy, pulmonary infection, and hypertension on November 6, 2012. Following admission, findings from laboratory tests were as follows: white blood cell count, 14.76 9 109/L; hemoglobin, 123 g/L; platelet count, 28 9 109/L; creatinine, 272 lmol/L; fasting glucose, 14.26 mmol/L; prothrombin time, 14.6 s; D-dimer, 4800 lg/L. Tests for antinuclear and antineutrophil cytoplasmic antibodies were negative, but were positive for platelet-associated IgG. Dipotassium EDTA, sodium citrate, and heparin sodium were selected as the anticoagulants for blood cell counts, to exclude the influence of EDTA salt. We performed whole-blood cell counts measured by a Sysmex XE-2100 complete blood count analyzer, followed by blood smears and staining with a Sysmex sp1000i automated hematology slide preparation unit. The platelet and white blood cell counts were 12 9 109 and 6.9 9 109/L for EDTA-2K anticoagulants, 242 9 109 and 4.3 9 109/L for sodium citrate anticoagulants, 250 9 109 and 4.6 9 109/L for heparin sodium anticoagulants, respectively. The blood smear results are shown in Figs. 1 and 2. The International Committee for Standardization in Hematology recommends dipotassium EDTA for anticoagulation of peripheral blood cells for analysis [1]. Blood specimens that met the review criteria after detection by the blood cell analyzer were smeared, stained, and observed under a microscope. When the platelet aggregation occurred, a cluster of platelets with uneven sizes was
W. Xu B. Chen Y. Chen (&) Department of Clinical Laboratory, The First Affiliated Hospital, School of Medicine, Zhejiang University, 79 Qingchun Road, Hangzhou 310003, Zhejiang, China e-mail: [email protected]
observed. However, the aggregated platelets retained their individual morphologies (Fig. 3). Microscopic examination of the blood smear revealed clumps of violescent material
Fig. 1 With dipotassium EDTA (EDTA-2K) as anticoagulant. Clumps of violescent material with uneven edges (black arrow) in which there are purple particles (red arrow) can be observed
Fig. 2 With sodium citrate or heparin sodium as anticoagulants. Platelets are dispersed, and the morphology is complete (indicated by black arrow)
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Fig. 3 With EDTA-2K as anticoagulant in typical cases. Clumps of platelets aggregated with complete morphology (indicated by black arrow)
and occasional purple particles. Further, complete morphology of platelets was not observed. We were unable to confirm platelet aggregation from morphology alone, but flow cytometric determination of platelet counts after CD45 and CD61 fluorescent antibody labeling confirmed that the clumps of violescent material were aggregated platelets, thereby avoiding the misdiagnosis of PTCP. The test results using the three anticoagulants indicated that this was a typical case of EDTA-dependent platelet aggregation, and that the results could be revised by other anticoagulants. Microscopic examination of the blood smear using other anticoagulants showed that the morphology of the platelets was complete and scattered. The white blood cell count results with the different anticoagulants indicated that the platelet aggregation had an effect on leukocyte count. It was challenging to accurately distinguish platelet aggregates consisting of nucleic acid from white blood cells using the blood cell analyzer, due to their similar volume.
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The reason for the platelet aggregation is unclear, but it is generally considered to be associated with the fibrinogen receptor, glycoprotein complex IIb/IIIa (GPIIb/IIIa). Indirect evidence suggests that PTCP often occurs in patients exposed to GPIIb/IIIa receptor antagonist [2]. Currently, the hypothesis is that the antigen and antibody binding sites are hidden within the GPIIb/IIIa complex under ordinary conditions, and that the structures of the binding sites are altered and platelet aggregation occurs in the presence of EDTA. In conclusion, when a low blood platelet count is noted in patients, a blood smear and microscopic examination should be performed following strict review criteria. Platelet morphology should also be considered, so that the presence of platelets is not missed when platelet aggregation is observed. Acknowledgments This study was supported by Education Department of Zhejiang province (Y201226248). Conflict of interest interests.
All the authors declare they have no conflict of
References 1. Recommendations of the International Council for Standardization in Haematology for Ethylenediaminetetraacetic acid anticoagulation of blood for blood cell counting and sizing. International Council for Standardization in Haematology: expert panel on cytometry. Am J Clin Pathol. 1993;100(4):371–372. 2. Dabadie M, Valli N, Jacobin MJ, Laroche-Traineau J, Barat JL, Ducassou D, Nurden AT, Clofent-Sanchez G. Characterisation, cloning and sequencing of a conformation-dependent monoclonal antibody to the alphaIIbbeta3 integrin: interest for use in thrombus detection. Platelets. 2001;12(7):395–405.
IMAGES IN HEMATOLOGY
Ethylenediaminetetraacetate-dependent pseudothrombocytopenia accompanied by morphological changes Weiyi Xu • Baode Chen • Yu Chen
Received: 25 December 2013 / Revised: 20 February 2014 / Accepted: 20 February 2014 Ó The Japanese Society of Hematology 2014
A 65-year-old female patient was referred to our hospital for diabetes mellitus, diabetic nephropathy, pulmonary infection, and hypertension on November 6, 2012. Following admission, findings from laboratory tests were as follows: white blood cell count, 14.76 9 109/L; hemoglobin, 123 g/L; platelet count, 28 9 109/L; creatinine, 272 lmol/L; fasting glucose, 14.26 mmol/L; prothrombin time, 14.6 s; D-dimer, 4800 lg/L. Tests for antinuclear and antineutrophil cytoplasmic antibodies were negative, but were positive for platelet-associated IgG. Dipotassium EDTA, sodium citrate, and heparin sodium were selected as the anticoagulants for blood cell counts, to exclude the influence of EDTA salt. We performed whole-blood cell counts measured by a Sysmex XE-2100 complete blood count analyzer, followed by blood smears and staining with a Sysmex sp1000i automated hematology slide preparation unit. The platelet and white blood cell counts were 12 9 109 and 6.9 9 109/L for EDTA-2K anticoagulants, 242 9 109 and 4.3 9 109/L for sodium citrate anticoagulants, 250 9 109 and 4.6 9 109/L for heparin sodium anticoagulants, respectively. The blood smear results are shown in Figs. 1 and 2. The International Committee for Standardization in Hematology recommends dipotassium EDTA for anticoagulation of peripheral blood cells for analysis [1]. Blood specimens that met the review criteria after detection by the blood cell analyzer were smeared, stained, and observed under a microscope. When the platelet aggregation occurred, a cluster of platelets with uneven sizes was
W. Xu B. Chen Y. Chen (&) Department of Clinical Laboratory, The First Affiliated Hospital, School of Medicine, Zhejiang University, 79 Qingchun Road, Hangzhou 310003, Zhejiang, China e-mail: [email protected]
observed. However, the aggregated platelets retained their individual morphologies (Fig. 3). Microscopic examination of the blood smear revealed clumps of violescent material
Fig. 1 With dipotassium EDTA (EDTA-2K) as anticoagulant. Clumps of violescent material with uneven edges (black arrow) in which there are purple particles (red arrow) can be observed
Fig. 2 With sodium citrate or heparin sodium as anticoagulants. Platelets are dispersed, and the morphology is complete (indicated by black arrow)
123
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Fig. 3 With EDTA-2K as anticoagulant in typical cases. Clumps of platelets aggregated with complete morphology (indicated by black arrow)
and occasional purple particles. Further, complete morphology of platelets was not observed. We were unable to confirm platelet aggregation from morphology alone, but flow cytometric determination of platelet counts after CD45 and CD61 fluorescent antibody labeling confirmed that the clumps of violescent material were aggregated platelets, thereby avoiding the misdiagnosis of PTCP. The test results using the three anticoagulants indicated that this was a typical case of EDTA-dependent platelet aggregation, and that the results could be revised by other anticoagulants. Microscopic examination of the blood smear using other anticoagulants showed that the morphology of the platelets was complete and scattered. The white blood cell count results with the different anticoagulants indicated that the platelet aggregation had an effect on leukocyte count. It was challenging to accurately distinguish platelet aggregates consisting of nucleic acid from white blood cells using the blood cell analyzer, due to their similar volume.
123
The reason for the platelet aggregation is unclear, but it is generally considered to be associated with the fibrinogen receptor, glycoprotein complex IIb/IIIa (GPIIb/IIIa). Indirect evidence suggests that PTCP often occurs in patients exposed to GPIIb/IIIa receptor antagonist [2]. Currently, the hypothesis is that the antigen and antibody binding sites are hidden within the GPIIb/IIIa complex under ordinary conditions, and that the structures of the binding sites are altered and platelet aggregation occurs in the presence of EDTA. In conclusion, when a low blood platelet count is noted in patients, a blood smear and microscopic examination should be performed following strict review criteria. Platelet morphology should also be considered, so that the presence of platelets is not missed when platelet aggregation is observed. Acknowledgments This study was supported by Education Department of Zhejiang province (Y201226248). Conflict of interest interests.
All the authors declare they have no conflict of
References 1. Recommendations of the International Council for Standardization in Haematology for Ethylenediaminetetraacetic acid anticoagulation of blood for blood cell counting and sizing. International Council for Standardization in Haematology: expert panel on cytometry. Am J Clin Pathol. 1993;100(4):371–372. 2. Dabadie M, Valli N, Jacobin MJ, Laroche-Traineau J, Barat JL, Ducassou D, Nurden AT, Clofent-Sanchez G. Characterisation, cloning and sequencing of a conformation-dependent monoclonal antibody to the alphaIIbbeta3 integrin: interest for use in thrombus detection. Platelets. 2001;12(7):395–405.
