How Much Does the Platelet Aggregate Influence the Total Leukocyte Count? Comparison Between Manual and Automated Impedance Methods in Domestic Cats

Leticia Gomes Zanfagnini, Siham Kassab, Diefrey Campos, Jéssica Karoline de Oliveira Chaves, Acácio Duarte Pacheco

Abstract


Background: Automated hematology analyzers have been developed to optimize the time between analyses and have promising precision and accuracy. Complete blood count (CBC) is often requested as part of veterinary clinical examination. Automated analyzers are often used to determine CBCs, since processing as well as container-related errors may occur owing to variable sizes, aggregates, white or red blood cell fragments, and effects of EDTA on cell morphology. Platelet aggregates frequently occur in felines, with studies reporting a prevalence of approximately 71%. The aim of the present study was to evaluate the influence of exercise aggregates on the global white blood cell count of domestic cats using automated hematological counters with the impedance method.

Materials, Methods & Results: Blood samples of 140 cats, irrespective of age, sex, and breed, were collected into EDTA-containing tubes. The samples were obtained via routine clinical examinations at the Veterinary Hospital of the Federal Rural University of Rio de Janeiro (UFRRJ) and processed at the Veterinary Parasitology Experimental Chemotherapy Laboratory (LQEPV), belonging to the same institution. All the samples were processed on the Sysmex pocH-100iV Diff automated hematology apparatus according to the manufacturer's recommendations. Leukocyte counts were also manually determined using a duplicate Neubauer chamber. Standard dilutions were prepared immediately after the automated analysis. To identify the occurrence of platelet aggregates, a blood smear was made and visualized under a brightfield microscope at a magnification of 10× and scored 0 to 3 (G1, G2, G3, and G4) based on the aggregation intensity. In case of changes, the groups were subdivided according to the intensity of occurrence. Of the 140 samples analyzed, 76.4% (107/140) showed some degree of platelet aggregation. The maximum variation in leukocyte counts determined by the automatic and the manual technique in G1 was 2,500 cells. In G2, it was possible to identify a variation of 6,500 nucleated cells, whereas in G3, this value was 7,100 cells. In G4, where platelet aggregation was intense, the variation between counts was up to 15,000 nucleated cells. A significant difference of variation in total white blood cell count between manual and automated methods was observed when compared to animals that did not show any degree of platelet aggregation (P < 0.05). Of the total samples, 23.57% (33/140) comprised G1, 24.28% (34/140) G2, 22.14% (31/140) G3, and 30% (42/140) G4. Of the 140 samples analyzed, 107 showed aggregates, pseudo-thrombocytopenia, and changes in the total number of leukocytes.

Discussion: Samples with higher platelet aggregate formation showed greater interference in global leukometry when analyzed using the hematological counter. White blood cell counts determined by automated analyzers should be interpreted with caution and compared to manual counts when there is significant platelet aggregation in the sample. The findings reinforce the importance of reconfirming the results obtained using an automated equipment in order to avoid misinterpretations that may influence diagnosis and therapy. It is essential to re-check the values obtained from an automated equipment with traditional methods in order to minimize possible errors generated by the equipment, since such errors may affect the clinical diagnosis and subsequently, the therapeutic approach chosen.


Full Text:

PDF

References


Ayres M., Ayres Jr. D. & Santos A. 2007. BioEstat 5.0. Imprensa Oficial do Estado do Pará. 323.

Bauer N., Nakagawa J., Dunker C., Failing K. & Moritz A. 2012. Evaluation of the automated hematology analyzer Sysmex XT-2000 i VTM compared to the ADVIA® 2120 for its use in dogs, cats, and horses. Part II: Accuracy of leukocyte differential and reticulocyte count, impact of anticoagulant and sample aging. Journal of Veterinary Diagnostic Investigation. 24(1): 74-89.

Becker M., Moritz A. & Giger U. 2008. Comparative clinical study of canine and feline total blood cell count results with seven in-clinic and two commercial laboratory hematology analyzers. Veterinary Clinical Pathology. 37(4): 373-384.

Bhar V.S. & Singh R. 2019. Platelet satellitism: unusual cause of spurious thrombocytopenia. Journal of Hematopathology. 12(3): 171-172.

Ghariani I., Braham N., Hamzaoui S. & Bekir L. 2017. Platelet satellitism in autoimmune hemolytic anemia. Current Research in Translational Medicine. 65(2): 61-64.

Granat F., Geffré A., Bourgès-Abella N., Braun J.P. & Trumel C. 2013. Changes in haematology measurements with the Sysmex XT-2000iV during storage of feline blood sampled in EDTA or EDTA plus CTAD. Journal of Feline Medicine and Surgery. 15(6): 433-444.

Kim S.Y., Kim J.E., Kim H.K., Han K.S. & Toh C.H. 2010. Accuracy of Platelet Counting by Automated Hematologic Analyzers in Acute Leukemia and Disseminated Intravascular Coagulation: Potential Effects of Platelet Activation. American Journal of Clinical Pathology. 134(4): 634-647.

Mezaroba M.E., Thomé J., Peres L.R.R., Rodrigues G. & Veiga A.P.M. 2018. New veterinary reference values for mean platelet volume (MVP), platelet distribution width (PDW) and platelet count (PCT) in the Curitibanos microregion. Revista Brasileira de Análises Clínicas. 50(2):171-173

Moritz A. & Hoffmann C. 1997. Platelet count in the cat. Tierarztliche Praxis. Ausgabe K, Kleintiere/Heimtiere. 25(6): 695-700.

Norman E.J., Barron R.C.J., Nash A.S. & Clampitt R.B. 2001. Evaluation of a Citrate-Based Anticoagulant with Platelet Inhibitory Activity for Feline Blood Cell Counts. Veterinary Clinical Pathology. 30(3): 124-132.

Rajajee S., Subbiah E., Krishnamurthy N., Paranjothi S. & Lohiya N. 2019. Pseudothrombocytopenia and Usefulness of Platelet Aggregates in Peripheral Smear in the Diagnosis of Scrub Typhus. The Indian Journal of Pediatrics. 86(1): 93-94.

Riond B., Waßmuth A.K., Hartnack S., Hofmann L.R. & Lutz H. 2015. Effective prevention of pseudothrombocytopenia in feline blood samples with the prostaglandin I2 analogue Iloprost. BMC Veterinary Research. 11:183-191.

Riond B., Weissenbacher S., Hofmann L. R. & Lutz H. 2011. Performance evaluation of the Sysmex pocH-100iV Diff hematology analyzer for analysis of canine, feline, equine, and bovine blood. Veterinary Clinical Pathology. 40(4): 484-495.

• 14 Sousa S.M., Sousa T.M., Silva C.F. & Mendes C.C. 2019. Pseudothrombocytopenia: a case of platelet satellitism and phagocytosis by neutrophils. Platelets. 1 (45): 1-3.

Tang W., Tang C., Zheng J., He Y. & Lu F. 2018. Fragmentation of Red Blood Cells Caused Pseudothrombocytosis in a Patient. Clinical Laboratory. 64(6): 1071-1074.

Tvedten H.W., Bäcklund K. & Lilliehöök I.E. 2015. Reducing error in feline platelet enumeration by addition of Iloprost to blood specimens: comparison to prostaglandin E1 and EDTA. Veterinary Clinical Pathology. 44(2): 179-187.

Zandecki M., Genevieve F., Gérard J. & Godon A. 2012. Spurious Counts and Spurious Results on Hematology Analyzers: White Blood Cells, Red Blood Cells, Hemoglobin, Red Cell Indices, and Reticulocytes. In: K. Kottke-Marchant & B.H. Davis (Eds). Laboratory Hematology Practice. Oxford: Wiley-Blackwell, pp.79-95.

Zelmanovic D. & Hetherington E.J. 1998. Automated Analysis of Feline Platelets in Whole Blood, Including Platelet Count, Mean Platelet Volume, and Activation State. Veterinary Clinical Pathology. 27(1): 2-9.

Table 1. Variation in the number of leukocytes, total from automatic and manual counting, in relation to platelet aggregate type.

Figure 1. Photomicrograph of the fringe region of the hematological extension of cats, showing the severity of platelet aggregation formation under optical microscopy at 10× magnification. A- Absence of platelet aggregate formation. B- Arrows showing mild platelet aggregate formation. C- Arrows showing moderate platelet aggregate formation. D- Arrows showing intense platelet aggregate formation.

Figure 2. Determination of total number variation of leukocytes in feline patients submitted to the impedance-automated leukocyte counting methods and manual counting, according to the presence of platelet aggregation.




DOI: https://doi.org/10.22456/1679-9216.105557

Copyright (c) 2021 Acácio Pacheco

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.