Figure 6.
Lactic acid treatment does not affect viability, yield and IL-3 induced activation of basophils, but removes only a small fraction of the cell surface bound IgE. Basophils were incubated on ice either with phosphate buffered saline (PBS) or ice-cold lactic acid buffer pH 3.9 (LA). Cells were then washed and cultured in serum-free X-VIVO 15 medium (0.1 × 106 cells/well per 200 µL) in 96-well U-bottomed plate along with IL-3 (100 ng/0.5 million cells) for 24 h. (A) The viability of cells immediately following LA treatment as analyzed by fixable viable dye staining. (B) The forward and side scatter plot of LA-treated basophils following 24-h culture in IL-3. (C) The expression of basophil activation markers ST2, FcεRI and CD69 in LA-treated cells after 24 h culture in IL-3 (mean ± SD, n = 4 experiments from two donors). (D) Efficacy of stripping of basophil surface-bound IgE by LA as analyzed by surface staining of IgE and analyses by flow cytometry. The intensity of IgE on the surface of basophils was represented by MFI values (Median fluorescence intensity). ** p < 0.01; *** p < 0.001; **** p < 0.0001; one-way ANOVA with Tukey’s multiple comparison test.