Abstract
The development of new advances in understanding the role of neutrophils in inflammation requires effective procedures for isolating and purifying neutrophils. Methods for isolating human neutrophils are fairly standard, and some are covered in other chapters of this volume and previous editions. However, procedures for isolating neutrophils from nonhuman species used to model human diseases vary from those used in isolating human neutrophils and are not as well developed. Since neutrophils are highly reactive and sensitive to small perturbations, the methods of isolation are important to avoid isolation technique-induced alterations in cell function. We present methods here for reproducibly isolating highly purified neutrophils from large animal models (bovine, equine, ovine), small animal models (murine and rabbit), and nonhuman primates (cynomolgus macaques) and describe optimized details for obtaining the highest cell purity, yield, and viability.
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References
Wiles S, Hanage WP, Frankel G et al (2006) Modelling infectious disease - time to think outside the box? Nat Rev Microbiol 4:307–312
Webb DR (2014) Animal models of human disease: inflammation. Biochem Pharmacol 87:121–130
Casal M, Haskins M (2006) Large animal models and gene therapy. Europ J Hum Genet 14:266–272
Ziegler A, Gonzalez L, Blikslager A (2016) Large animal models: the key to translational discovery in digestive disease research. Cell Mol Gastroenterol Hepatol 2:716–724
Styrt B (1989) Species variation in neutrophil biochemistry and function. J Leukoc Biol 46:63–74
Glasser L, Fiederlein RL (1990) The effect of various cell separation procedures on assays of neutrophil function. A critical appraisal. Am J Clin Pathol 93:662–669
Watson F, Robinson JJ, Edwards SW (1992) Neutrophil function in whole blood and after purification - changes in receptor expression, oxidase activity and responsiveness to cytokines. Biosci Rep 12:123–133
Forsyth KD, Levinsky RJ (1990) Preparative procedures of cooling and re-warming increase leukocyte integrin expression and function on neutrophils. J Immunol Methods 128:159–163
Macey MG, Jiang XP, Veys P et al (1992) Expression of functional antigens on neutrophils. Effects of preparation. J Immunol Methods 149:37–42
Alvarez-Larrán A, Toll T, Rives S et al (2005) Assessment of neutrophil activation in whole blood by flow cytometry. Clin Lab Haematol 27:41–46
Pycock JF, Allen WE, Morris TH (1987) Rapid, single-step isolation of equine neutrophils on a discontinuous percoll density gradient. Res Vet Sci 42:411–412
Lowell CA, Fumagalli L, Berton G (1996) Deficiency of Src family kinases p59/61hck and p58c-fgr results in defective adhesion-dependent neutrophil functions. J Cell Biol 133:895–910
Woldehiwet Z, Scaife H, Hart CA et al (2003) Purification of ovine neutrophils and eosinophils: anaplasma phagocytophilum affects neutrophil density. J Comp Pathol 128:277–282
White-Owen C, Alexander JW, Sramkoski RM et al (1992) Rapid whole-blood microassay using flow cytometry for measuring neutrophil phagocytosis. J Clin Microbiol 30:2071–2076
Doerschuk CM, Allard MF, Martin BA et al (1987) Marginated pool of neutrophils in rabbit lungs. J Appl Physiol 63:1806–1815
Swamydas M, Lionakis MS (2013) Isolation, purification and labeling of mouse bone marrow neutrophils for functional studies and adoptive transfer experiments. J Vis Exp (77):e50586
Boxio R, Bossenmeyer-Pourié C, Steinckwich N et al (2004) Mouse bone marrow contains large numbers of functionally competent neutrophils. J Leukoc Biol 75:604–611
Siemsen DW, Malachowa N, Schepetkin IA et al (2014) Neutrophil isolation from nonhuman species. Methods Mol Biol 1124:19–37
DeLeo FR, Renee J, Mccormick S et al (1998) Neutrophils exposed to bacterial lipopolysaccharide upregulate NADPH oxidase assembly. J Clin Invest 101:455–463
Acknowledgments
This work was supported in part by National Institutes of Health IDeA Program COBRE Grant GM110732; the Intramural Research Program of the National Institute of Allergy and Infectious Diseases, National Institutes of Health; USDA National Institute of Food and Agriculture Hatch project 1009546; and the Montana State University Agricultural Experiment Station.
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Siemsen, D.W. et al. (2020). Isolation of Neutrophils from Nonhuman Species. In: Quinn, M., DeLeo, F. (eds) Neutrophil. Methods in Molecular Biology, vol 2087. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0154-9_4
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DOI: https://doi.org/10.1007/978-1-0716-0154-9_4
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