Single‐step enrichment of basophils from human peripheral blood by a novel method using a Percoll density gradient |
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| Authors: | Hiroyuki Shiono Takuya Matsui Tadashi Okada Yoichiro Ito |
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| Institution: | 1. Department of Physiology, Aichi Medical University School of Medicine, Nagakute‐city, Aichi, Japan;2. Bioseparation Technology Laboratory, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD, USA |
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| Abstract: | 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 × 105 cells in fraction 3 and 1.61 × 105 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. |
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| Keywords: | Basophils Cell separation Centrifugation Density gradient Human peripheral blood |
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