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J. Dairy Sci. 86:828-834
© American Dairy Science Association, 2003.

Differential Leukocyte Count Method for Bovine Low Somatic Cell Count Milk

H. Dosogne, F. Vangroenweghe, J. Mehrzad, A. M. Massart-Leën and C. Burvenich

Ghent University, Faculty of Veterinary Medicine, Department of Physiology, Biochemistry and Biometrics Salisburylaan 133, 9820 Merelbeke, Belgium

Corresponding author:
C. Burvenich; e-mail:
christian.burvenich{at}rug.ac.be.

Whereas many differential leukocyte count methods for high somatic cell count (SCC) milk from mastitic cows are available, only a few have been developed for low SCC milk. We have developed a flow cytometric differential leukocyte count method for low SCC milk. The procedure consists of 1) 1.5 ml of diluted milk sample (30%, vol/vol dilution with PBS), 2) centrifugation, 3) leukocyte labeling with SYTO 13 and 4) flow cytometric analysis. Four major leukocyte populations can be clearly identified in the green fluorescence-side scatter dot plot: lymphocytes and monocytes (LM), polymorphonuclear neutrophils (PMN), mature macrophages (M{Phi}), and cells with apoptotic features based on chromatin condensation and nuclear fragmentation. The optimal processing temperature was 20°C. Significant differences among samples with similar differential leukocyte counts were found. Storage of milk samples during 2 d at 7°C had no effect on differential leukocyte count. Using the new method, differential leukocyte count was performed in low SCC milk samples from cows in early, mid, and late lactation. In accordance with previous studies, PMN and M{Phi} percentages were lower and LM percentages were higher in early lactation than in the other stages of lactation. The percentage of cells with apoptotic features was higher in early lactation than in mid and late lactation. In conclusion, a rapid, simple, accurate, and reproducible standard procedure was developed to determine the differential leukocyte count (M{Phi}, PMN, LM, and cells with apoptotic features) of bovine low SCC milk.

Key Words: differential leukocyte count • flow cytometry • low somatic cell count milk

Abbreviation key: FS = forward scatter, LM = lymphocytes and monocytes, M{Phi} = macrophage, PMN = polymorphonuclear neutrophil, SSC = side scatter




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J Dairy Sci, October 1, 2006; 89(10): 4066 - 4069.
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