Abstract
Cell-free granules, upon extrusion from human eosinophils, remain fully competent to secrete granule-derived proteins in receptor-mediated processes in response to different stimuli. However, in order to avoid the shrinkage and damage of granules, as well as preserve their structure, properties, and functionality, the use of an optimized process of subcellular fractionation using an isoosmotic density gradient is needed. Here, we describe a detailed protocol of subcellular fractionation of nitrogen-cavitated eosinophils on an isoosmotic iodinated density gradient (iodixanol) and the isolation of well-preserved and functional membrane-bound specific granules.
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Acknowledgment
The authors thank Conselho Nacional de Pesquisas (CNPq—Brazil), Comissão de Aperfeiçoamento de NÃvel Superior (Capes—Brazil), and Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ—Brazil) for the financial support.
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Baptista-dos-Reis, R., Muniz, V.S., Neves, J.S. (2014). Isolation and Functional Assessment of Eosinophil Crystalloid Granules. In: Walsh, G. (eds) Eosinophils. Methods in Molecular Biology, vol 1178. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1016-8_9
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DOI: https://doi.org/10.1007/978-1-4939-1016-8_9
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Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-1015-1
Online ISBN: 978-1-4939-1016-8
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