Pre-analytical and analytical variability of reticulocyte counts in dogs.
automated reticulocyte enumeration
manual count
reticulocytosis
staining artefacts
storage effect
Journal
The Veterinary record
ISSN: 2042-7670
Titre abrégé: Vet Rec
Pays: England
ID NLM: 0031164
Informations de publication
Date de publication:
06 May 2023
06 May 2023
Historique:
revised:
28
01
2023
received:
04
07
2022
accepted:
06
02
2023
medline:
8
6
2023
pubmed:
14
3
2023
entrez:
13
3
2023
Statut:
ppublish
Résumé
Automated fluorescence-based haematology analysers are now available for reticulocyte enumeration in veterinary medicine, but manual counting is still largely used. This study aimed to evaluate potential sources of analytical and pre-analytical errors when performing automated and manual counts. Automated and two-operator double-blind manual reticulocyte counts were performed on 15 blood samples. The intra-assay variation of the automated and manual counts and the interoperator variation in the manual counts were then calculated. In addition, the effects of storage were evaluated using samples refrigerated at 4°C or stored at room temperature for 2, 4, 12, 24, 48 or 72 hours after sampling. Intra-assay coefficients of variation were lower for automated counts than for manual counts. Comparison between automated and mean total manual reticulocyte count showed no significant differences. In both refrigerated samples and those stored at room temperature, an increase in reticulocyte count was recorded only after 72 hours. Staining artefacts occurred only in one stored sample counted manually. The presence of cytoplasmic particles other than RNA can cause misinterpretation of cells, leading to an erroneous reticulocyte count. The use of an automated analyser is preferable for reticulocyte enumeration in dogs. Common storage conditions seem to minimally affect reticulocyte evaluation; however, it is recommended to perform the analysis as soon as possible after sampling.
Sections du résumé
BACKGROUND
BACKGROUND
Automated fluorescence-based haematology analysers are now available for reticulocyte enumeration in veterinary medicine, but manual counting is still largely used. This study aimed to evaluate potential sources of analytical and pre-analytical errors when performing automated and manual counts.
METHODS
METHODS
Automated and two-operator double-blind manual reticulocyte counts were performed on 15 blood samples. The intra-assay variation of the automated and manual counts and the interoperator variation in the manual counts were then calculated. In addition, the effects of storage were evaluated using samples refrigerated at 4°C or stored at room temperature for 2, 4, 12, 24, 48 or 72 hours after sampling.
RESULTS
RESULTS
Intra-assay coefficients of variation were lower for automated counts than for manual counts. Comparison between automated and mean total manual reticulocyte count showed no significant differences. In both refrigerated samples and those stored at room temperature, an increase in reticulocyte count was recorded only after 72 hours. Staining artefacts occurred only in one stored sample counted manually.
LIMITATIONS
CONCLUSIONS
The presence of cytoplasmic particles other than RNA can cause misinterpretation of cells, leading to an erroneous reticulocyte count.
CONCLUSION
CONCLUSIONS
The use of an automated analyser is preferable for reticulocyte enumeration in dogs. Common storage conditions seem to minimally affect reticulocyte evaluation; however, it is recommended to perform the analysis as soon as possible after sampling.
Types de publication
Randomized Controlled Trial, Veterinary
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
e2733Informations de copyright
© 2023 British Veterinary Association.
Références
Tvedten HW, Moritz A. Reticulocytes and Heinz body staining and enumeration. In: Weiss DJ, Wardrop KJ, editors. Schalm's veterinary hematology. 6th ed. Ames, IA: Wiley-Blackwell; 2010. p. 1067-73.
Riley RS, Ben-Ezra JM, Tidwell A, Romagnoli G. Reticulocyte analysis by flow cytometry and other techniques. Hematol Oncol Clin North Am. 2002;16:373-420.
Cowgill ES, Neel JA, Grindem CB. Clinical application of reticulocyte counts in dogs and cats. Vet Clin North Am Small Anim Pract. 2003;33:1223-44.
George L, Basu D, Kar R. Comparison between manual and automated methods of counting reticulocytes and the effect of sample storage on reticulocyte count: a cross-sectional study from Southern India. Indian J Hematol Blood Transfus. 2022;38(1):106-10.
Gilmer PR, Koepke JA. The reticulocyte. An approach to definition. Am J Clin Pathol. 1976;66:262-7.
Peebles D, Hochberg A, Clark T. Analysis of manual reticulocyte counting. Am J Clin Pathol. 1981;76:713.
NCCLS. Methods for Reticulocyte Counting (Automated Blood Cell Counters, Flow Cytometry, and Supravital Dyes); Approved Guideline-Second Edition. NCCLS document H44-A2 [ISBN 1-56238-527-5], Vol. 24 No. 8, 2004.
Imeri F, Herklotz R, Risch L, Arbetsleitner C, Zerlauth M, Risch GM, et al. Stability of hematological analytes depends on the hematology analyser used: a stability study with Bayer Advia 120, Beckman Coulter LH 750 and Sysmex XE 2100. Clin Chim Acta. 2008;397:68-71.
Bourgès-Abella NH, Geffré A, Deshuillers PL, Braun J-PD, Trumel C. Changes in hematology measurements in healthy and diseased dog blood stored at room temperature for 24 and 48 hours using the XT-2000iV analyzer. Vet Clin Pathol. 2014;43(1):24-35.
Granat F, Geffré A, Bourgès-Abella N, Braun J-P, Trumel C. Changes in haematology measurements with the Sysmex XT-2000iV during storage of feline blood sampled in EDTA or EDTA plus CTAD. J Feline Med Surg. 2013;15(6):433-44.
Layssol-Lamour C, Lavabre T, Braun JP, Trumel C, Bourgès-Abella N. The effects of storage at 4°C and 20°C on the hemograms of C57BL/6 mice and Wistar rats using the IDEXX ProCyte Dx and blood smear evaluations. Vet Clin Pathol. 2019;48(4):652-67.
Fujino Y, Nakamura Y, Matsumoto H, Fukushima K, Takahashi M, Ohno K, et al. Development and evaluation of a novel in-clinic automated hematology analyzer, ProCyte Dx, for canine erythrocyte indices, leukogram, platelet counts and reticulocyte counts. J Vet Med Sci. 2013;75(11):1519-24.
Christian JA. Erythrokinetics and erythrocyte destruction. In: Weiss DJ, Wardrop KJ, editors. Schalm's veterinary hematology. 6th ed. Ames, IA: Wiley-Blackwell; 2010. p. 136-43.
Paltrinieri S, Rossi G, Manca M, Scarpa P, Vitiello T, Giordano A. Sensitivity and specificity of manual and automated measurements of reticulocyte parameters for classification of anemia in dogs: 174 cases (1993-2013). J Am Vet Med Assoc. 2016;249(7):776-86.
Wenger-Riggenbach B, Hässig M, Hofmann-Lehmann R, Lutz H. Evaluation of the LaserCyte: an in-house hematology analyzer for dogs and cats. Comp Clin Pathol. 2006;15:117-29.
Nabity MB, Harr KE, Camus MS, Flatland B, Vap LM. ASVCP guidelines: allowable total error hematology. Vet Clin Pathol. 2018;53:833-9.
Westgard JO. Method validation. In: Westgard JO, editor. Basic method validation. 2nd ed. Madison, WI: Westgard QC; 2003. p. 156-7.
Savage RA, Skoog DP, Rabinovitch A. Analytic inaccuracy and imprecision in reticulocyte counting: a preliminary report from the College of American Pathologists Reticulocyte Project. Blood Cells. 1985;11:97-112.
Riley RS, Ben-Ezra JM, Goel R, Tidwell A. Reticulocytes and reticulocyte enumeration. J Clin Lab Anal. 2001;15:267-94.
Grebert M, Granat F, Braun J-P, Leroy Q, Bourgès-Abella N, Trumel C. Validation of the Sysmex XN-V hematology analyzer for canine specimens. Vet Clin Pathol. 2021;50:184-97.
Bauer N, Nakagawa J, Dunker C, Failing K, Moritz A. Evaluation of the automated hematology analyzer Sysmex XT-2000iV™ 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. J Vet Diagn Invest. 2012;24:74-89.
Keiner M, Fuchs J, Bauer N, Moritz A. Evaluation of reticulocyte hemoglobin content (RETIC-HGB) for the diagnosis of iron-limited erythropoiesis in cats. Vet Clin Pathol. 2020;49:557-66.
Pinter E, László K, Schüszler I, Konderák J. The stability of quantitative blood count parameters using the ADVIA 2120i hematology analyzer. Pract Lab Med. 2015;4:16-21.
de Baca ME, Gulati G, Kocher W, Schwarting R. Effects of storage of blood at room temperature on hematologic parameters measured on Sysmex XE2100. Lab Med. 2006;37:28-35.
Seo JY, Lee ST, Kim SH. Performance evaluation of the new hematology analyzer Sysmex XN-series. Int J Lab Hematol. 2015;37:155-64.
Weiss D, Tvedten H. Erythrocyte disorders. In: Willard MD, Tvedten H, editors. Small animal clinical diagnosis by laboratory methods. 4th ed. St. Louis, MO: Elsevier Saunders; 2004. p. 38-62.