Eosinophil chemotaxis by chemokines: a study by a simple photometric assay.

Abstract

The effects of a panel of 15 chemokines on eosinophil chemotaxis were studied by a new photometric assay which is both less tedious and less laborious than the conventional manual counting methods. Approximately 40 chemokines have been identified to date, but there is little information on the eosinophil migration-inducing ability of chemokines other than CC chemokine receptor (CCR) 3 ligands. Eosinophil migration was measured by the Boyden chamber technique with a 96-well multiwell chamber and polycarbonate membrane filter. Eosinophil migration was assessed by determination of the eosinophil peroxidase (EPO) activity, and photometric measurement was performed with a microtiter plate reader. The assay was sensitive enough to detect 200 eosinophils, and the time required was within 4 h. CCR3 ligands, i.e., regulated on activation normal T-cell expressed and secreted (RANTES), eotaxin, eotaxin-2, and monocyte chemoattractant protein (MCP)-3, induced significant migration, while other chemokines showed no significant migration-inducing ability. Although the chemotaxis induction by these chemokines was efficiently inhibited by anti-CCR3 mAb, anti-CCR1 mAb failed to show any inhibitory effects. The photometric assay is suitable for analyzing a large number of samples. CCR3 ligands are the most important chemokines inducing eosinophil chemotaxis; thus, CCR3 represents a possible therapeutic target for the treatment of allergic diseases.