Efficient purification of eosinophils from human tissues: a comparative study.
BACKGROUND
Eosinophils are key effector cells in allergy and in other inflammatory diseases. Although they carry out their function in the tissues, no efficient method exists allowing for consistent purification of tissue eosinophils for culture. Rather, studies rely mainly on peripheral blood eosinophils. This study aimed to determine the most efficient protocol for purifying eosinophils from nasal polyp tissue.


METHODS
Nasal polyps were obtained from patients undergoing surgical polypectomy. The polyps were minced and enzymatically digested. Surface receptor analysis was performed by flow cytometry. In order to obtain optimal purification, the nasal polyp cell suspension was subjected to two methods of purification: 1) positive magnetic selection of CCR3+cells, or 2) negative selection using CD3/CD14/CD16 magnetic beads. Enriched tissue eosinophils were cultured with or without IL-3, IL-5 or GM-CSF, and their survival was evaluated by flow cytometry.


RESULTS
Tissue-derived eosinophils exhibited surface expression of NEC2, DNAM-1, NTBa, 2B4, and CD300a comparable to similarly prepared eosinophils obtained from the peripheral blood of the same patients. Positive selection consistently yielded eosinophils of high purity (>90%) with 63% viability. In contrast, negative selection yielded better viability (88%), reduced purity (66%), and could be utilized for in vitro activation experiments.


CONCLUSIONS
Eosinophils can be purified from nasal polyps. Negative selection appears to be advantageous due to improved viability of the eosinophils, which may be cultured and activated in vitro. This methodology is an important advance in studying tissue eosinophils for further investigations on inflammatory tissue responses.
Nissim Ben Efraim
A H
AH
Department of Pharmacology and Experimental Therapeutics, School of Pharmacy, Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, Israel.
Munitz
A
A
Sherman
Y
Y
Mazer
B D
BD
Levi-Schaffer
F
F
Eliashar
R
R
D002469:Cell Separation / Q000379:methods*; D002470:Cell Survival; D002478:Cells, Cultured; D004804:Eosinophils / Q000166:cytology* / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D008280:Magnetics; D009298:Nasal Polyps / Q000276:immunology*; D011956:Receptors, Cell Surface / Q000032:analysis
D011956:Receptors, Cell Surface