(A) Chemotactic activities of the recombinant CKβ8-1 and CKβ8. Lymphocytes, monocytes, and neutrophils were isolated from human peripheral blood buffy coat. Chemotaxis assays were performed using a Boyden chamber kit (Neuro Probe) with varying concentrations of CKβ8-1 and CKβ8. Microscopic observation at 100 × was used for counting each cell type. Three independent regions were selected and cell numbers were counted. The chemotactic index was driven by the following formula: (the number of cells chemoattracted by each concentration of CKβ8-1 or CKβ8)/(the number of cells chemoattracted by medium alone). (B) Each leukocyte population was isolated and examined for its response to CKβ8-1 and CKβ8, along with RANTES and MIP-1α. Two million cells were loaded with fura-2AM (Molecular Probes, Inc, Eugene, OR), stimulated with 25 nmol/L each chemokine, and used for Ca2+ flux assay.
