Mast cell activation and generation of supernatant samples
After 8 weeks of culture, IgE sensitized mast cells were activated by challenge with recombinant Der p 2 allergen in log10dilutions for 30 minutes in a water bath at 37° C. Activation was assessed by flow cytometry as surface expression of CD63. Supernatants were generated by activating MCs for 3, 6 or 24 hours at 37° C with an optimal concentration of Der p 2 allergen determined above. After centrifugation for 10 minutes, 4° C, 2000G, the supernatant and pellet were frozen separately. A supernatant sample not activated with allergen (time 0 hours) was saved as baseline control.