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Single‐step enrichment of basophils from human peripheral blood by a novel method using a Percoll density gradient
Author(s) -
Shiono Hiroyuki,
Matsui Takuya,
Okada Tadashi,
Ito Yoichiro
Publication year - 2016
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.201600329
Subject(s) - percoll , basophil , chromatography , chemistry , differential centrifugation , density gradient , flow cytometry , histamine , centrifugation , fractionation , immunology , microbiology and biotechnology , biology , antibody , immunoglobulin e , biochemistry , endocrinology , physics , quantum mechanics
We have developed a novel continuous flow‐through cell separation method using a Percoll density gradient. This method can continuously separate a large number of cells into five fractions according to their densities. To apply this method to the separation of basophils, Percoll density gradients were modified to improve basophil enrichment. When a set of Percoll density gradients was prepared (1.071, 1.075, 1.080, 1.084, and 1.090 g/mL) the basophils in a healthy volunteer were enriched by an average of 23.1 and 63.5% at Percoll densities of 1.075 (fraction 3) and 1.080 g/mL (fraction 4), respectively. On average, the yield of basophils was 1.66 × 10 5 cells in fraction 3 and 1.61 × 10 5 cells in fraction 4 from 9 mL of peripheral blood. The expression of CD203c (cluster of differentiation 203c) on separated basophils was upregulated by anti‐immunoglobulin E stimulation similar to basophils in whole blood. Histamine release induced by calcium ionophore was also observed in the separated basophils. The present method will be useful for basophil enrichment since it preserves their function without using counterflow elutriation and immunological reagents, and this method will be effective as a preparative separation for cell purification by flow cytometry.