Fig. 2

Neutralizing effect of patient plasma on GM-CSF-induced STAT5 phosphorylation: IC₅₀ A and EC₅₀ (B). PBMCs from the same healthy donor were cultured with plasma from patients and stimulated with recombinant human GM-CSF. After 30 min at 37 °C, cells were fixed and permeabilized for intracellular pSTAT5 staining in CD14 positive cells. A IC₅₀ experiments: The dilutions of patient plasma varied from 0.01 to 30% (diluted in AB serum) and the concentration of GM-CSF stayed constant (10 ng/ml). The dilution at which 50% inhibition of the GM-CSF-induced pSTAT5 occurred was determined for each plasma. B EC₅₀ experiments: The concentration of antibodies was normalized to 10 MMAU for each sample whereas the concentration of GM-CSF for stimulation varied ween 10^–1 to 10⁴ ng/ml. The concentration of GM-CSF responsible for 50% of maximal GM-CSF-induced pSTAT5 in presence of 10 MMAU of anti-GM-CSF antibody was assessed for each sample. Results for groups of patients with distinct phenotypes are expressed as mean ± SD and comparisons between groups were done using nonparametric t-test