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Nat Protoc. 2006;1(6):2613-20.

Highly purified selective isolation of eosinophils from human peripheral blood by negative immunomagnetic selection.

Author information

1
Section of Pulmonary and Critical Care Medicine, Department of Medicine and Pediatrics, The University of Chicago, Chicago, Illinois 60637, USA.

Abstract

Eosinophils are a minority constituent in human peripheral blood. The study of eosinophils has been limited by difficulty in achieving sufficient cell number and purity. We describe a modified protocol for immunomagnetic cell separation for efficient isolation of human peripheral blood eosinophils. We employ a mixture of monoclonal antibodies (mAbs) directed against cell-surface antigens on human hematopoietic cells combined with secondary labeling with a colloidal suspension of magnetic dextran-iron particles for negative selection of eosinophils. Unwanted labeled cells are retained in the magnetized column, permitting high recovery (70%) and purity (>98%) of eosinophils while retaining cell viability. Eosinophils remain quiescent after isolation, and stimulation caused by cytokines upregulates (i) cell-cell or cell-extracellular matrix protein adhesion, (ii) secretion of bioactive mediators and (iii) cell-surface adhesion molecules. This method for purified isolation is accomplished in < or = 4 h and preserves eosinophils in a quiescent, viable state.

PMID:
17406516
DOI:
10.1038/nprot.2006.340
[Indexed for MEDLINE]

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