Fig. 3. Varying extracellular pH predictably affects eosinophil depolarization. (A) Eosinophil intracellular pH was determined using BCECF/AM and calibrated as described in Materials and Methods. Buffer pH was verified and adjusted before each experiment. Unstimulated eosinophils were resuspended at 37°C at pH 6.0 (n=8), pH 6.5 (n=8), pH 7.0 (n=8) and pH 7.4 (n=8). The mean is stated above each bar (mean±s.e.m.). (B) Superoxide production was determined using the cytochrome c colorimetric assay as described in Materials and Methods. Buffer pH was verified and adjusted before each experiment. Cells were stimulated with 2 nM PMA at 37°C at pH 6.0 (n=5), pH 6.5 (n=5), pH 7.0 (n=5) and pH 7.4 (n=5). NADPH oxidase rate was calculated as described in Materials and Methods (means±s.e.m.). **P<0.01. (C) Eosinophil membrane potential was measured using diBAC₄(3) and flow cytometry. Media pH was verified and adjusted before each experiment. Eosinophils were stimulated with 800 nM at room temperature at pH 6.0 (; n=3), pH 6.5 (; n=4), pH 7.0 (; n=8), pH 7.4 (; n=8) and pH 8.0 (*; n=4). Means±s.e.m. are shown.

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