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

Background: The effects of a panel of 15 chemokines on eosinophil chemotaxi s were studied by a new photometric assay which is both less tedious and le ss laborious than the conventional manual counting methods. Approximately 4 0 chemokines have been identified to date, but there is little information on the eosinophil migration-inducing ability of chemokines other than CC ch emokine receptor (CCR) 3 ligands. Methods: Eosinophil migration was measured by the Boyden chamber technique with a 96-well multiwell chamber and polycarbonate membrane filter. Eosinop hil migration was assessed by determination of the eosinophil peroxidase (E PO) activity, and photometric measurement was performed with a microtiter p late reader. Results: The assay was sensitive enough to detect 200 eosinophils, and the time required was within 4 h. CCR3 ligands, i.e., regulated on activation n ormal T-cell expressed and secreted (RANTES), eotaxin, eotaxin-2, and monoc yte chemoattractant protein (MCP)-3, induced significant migration, while o ther chemokines showed no significant migration-inducing ability. Although the chemotaxis induction by these chemokines was efficiently inhibited by a nti-CCR3 mAb, anti-CCR1 mAb failed to show any inhibitory effects. Conclusions: The photometric assay is suitable for analyzing a large number of samples. CCR3 ligands are the most important chemokines inducing eosino phil chemotaxis; thus, CCR3 represents a possible therapeutic target for th e treatment of allergic diseases.