Stefan Dunzendorfer, MDa, Nicole C

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Functional expression of chemokine receptor 2 by normal human eosinophils Stefan Dunzendorfer, MDa, Nicole C. Kaneider, MDa, Arthur Kaser, MDb, Ewald Woell, MDa, José M.R. Frade, MDc, Mario Mellado, MDc, Carlos Martínez-Alonso, MDc, Christian J. Wiedermann, MDa Journal of Allergy and Clinical Immunology Volume 108, Issue 4, Pages 581-587 (October 2001) DOI: 10.1067/mai.2001.118518 Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 1 Dose response of eosinophils to various chemoattractants. Human eosinophils migrated into nitrocellulose filters toward soluble chemoattractants in the lower well of a Boyden microchemotaxis chamber. After fixing and staining, migration depth was quantified microscopically in the leading front assay. Data are expressed as means ± SEMs of the chemotaxis index (n = 5; see Materials and Methods section). Statistics: Mann-Whitney U test after Kruskal Wallis ANOVA (P < .001). n.s., Not significant. *P < .05; **P < .01 vs medium control. Journal of Allergy and Clinical Immunology 2001 108, 581-587DOI: (10.1067/mai.2001.118518) Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 2 RT-PCR analysis. Reverse transcriptase reaction was performed on 1 μg of RNA derived from 2 eosinophil preparations from healthy donors (E1, E2) and from monocytes (M) . As expected, size determination after gel electrophoresis revealed a single band of approximately 700 bp. Journal of Allergy and Clinical Immunology 2001 108, 581-587DOI: (10.1067/mai.2001.118518) Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 3 Cytofluorometry. CCR2 protein cell surface expression on eosinophils. Monocytes served as control. Solid lines indicate specific CCR2 mAb; bold lines indicate isotype-matched control. Data are representative of at least 3 independent experiments. Journal of Allergy and Clinical Immunology 2001 108, 581-587DOI: (10.1067/mai.2001.118518) Copyright © 2001 Mosby, Inc. Terms and Conditions

Fig. 4 Effect of MCP-1 on eosinophil chemotaxis. Human eosinophils alone or preincubated with a specific anti-CCR1, anti-CCR2, or anti-CCR3 blocking mAb migrated for 60 minutes toward various concentrations of human MCP-1 in the lower well of a Boyden microchemotaxis chamber. MCP-3 (0.1 nmol/L) served as positive control. After fixing and staining, migration depth was quantified microscopically in the leading front assay. Data are expressed as means ± SEMs of the chemotaxis index; n = 5. Statistics: Mann-Whitney U test after Kruskal Wallis ANOVA (P < .001). n.s., Not significant vs migration without mAb. #P < .05 vs migration with mAb. *P < .05; **P < .01 vs medium control. Journal of Allergy and Clinical Immunology 2001 108, 581-587DOI: (10.1067/mai.2001.118518) Copyright © 2001 Mosby, Inc. Terms and Conditions