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Flow cytometric analysis of intracellular complexity and CD45 expression for use in rapid differentiation of leukocytes in bovine blood samples

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  • 1 Department of Microbiology, Immunology, and Preventive Medicine, College of Veterinary Medicine, Iowa State University, Ames, IA 50011.
  • | 2 Periparturient Diseases of Cattle Research Unit, National Animal Disease Center, USDA-Agricultural Research Service, Ames, IA 50010.
  • | 3 Periparturient Diseases of Cattle Research Unit, National Animal Disease Center, USDA-Agricultural Research Service, Ames, IA 50010.
  • | 4 Present address is Pfizer Global Research and Development, Veterinary Medicine Pharmaceutical Discovery, 9303 S Carlisle St, Terre Haute, IN 47808.

Abstract

Objective—To develop an efficient and reliable method that accurately differentiates bovine lymphocytes from monocytes in leukograms.

Sample Population—Blood samples from 30 healthy cows and 1 calf with bovine leukocyte adhesion deficiency.

Procedure—Flow cytometric analysis of intracellular complexity and CD45 expression on bovine leukocytes was compared with results for conventional light microscopy methods. Verification of leukocyte subpopulations determined by intracellular complexity and CD45 expression was conducted, using 2-color phenotypic analysis with selected monoclonal antibodies.

Results—The CD45 and side-scatter properties of bovine leukocytes clearly differentiated cell types, including neutrophils, eosinophils, monocytes, and lymphocytes.

Conclusions and Clinical Relevance—This is a rapid assay that is simple to use. More importantly, it is more accurate than the conventional method that involves the use of blood slides and light microscopy, because of the ability of the assay to readily distinguish bovine monocytes and lymphocytes. Rapid preparation of samples and short analysis times allow for efficient and reliable examination of a large number of samples, and the task of viewing slides by light microscopy is eliminated. The labor-savings benefit of this procedure is most apparent in research environments that require frequent processing of batches of blood samples. (Am J Vet Res 2001;62:1740–1744)

Abstract

Objective—To develop an efficient and reliable method that accurately differentiates bovine lymphocytes from monocytes in leukograms.

Sample Population—Blood samples from 30 healthy cows and 1 calf with bovine leukocyte adhesion deficiency.

Procedure—Flow cytometric analysis of intracellular complexity and CD45 expression on bovine leukocytes was compared with results for conventional light microscopy methods. Verification of leukocyte subpopulations determined by intracellular complexity and CD45 expression was conducted, using 2-color phenotypic analysis with selected monoclonal antibodies.

Results—The CD45 and side-scatter properties of bovine leukocytes clearly differentiated cell types, including neutrophils, eosinophils, monocytes, and lymphocytes.

Conclusions and Clinical Relevance—This is a rapid assay that is simple to use. More importantly, it is more accurate than the conventional method that involves the use of blood slides and light microscopy, because of the ability of the assay to readily distinguish bovine monocytes and lymphocytes. Rapid preparation of samples and short analysis times allow for efficient and reliable examination of a large number of samples, and the task of viewing slides by light microscopy is eliminated. The labor-savings benefit of this procedure is most apparent in research environments that require frequent processing of batches of blood samples. (Am J Vet Res 2001;62:1740–1744)