Upregulation of CD23 by IL-4 and GM-CSF. Dose-ranging studies were performed to determine the optimum concentrations of IL-4 and GM-CSF. Alpha-smooth muscle isoactin positive Human ASMC (Clonetics) in T-75 flasks were starved for 24 h in 0.1% FBS containing medium M199. The cells were then stimulated with BSA (1 μg/ml), IL-4 (0.125. 0.25, 0.5, or 1.0 nM) or GM-CSF (0.1, 0.2, 0.4, or 0.8 nM) for 24 h. The cell lysates in RIPA buffer were subjected to western blot analysis for CD23. Mouse anti-human CD23 monoclonal antibody (clone M-L233, BD Biosciences, 1 μg/5 ml) was used as the primary antibody and anti-mouse horseradish peroxidase linked antibody as the secondary antibody (Amersham). The immunoreactive protein bands were detected by enhanced chemiluminescence light (ECL) (Amersham).
Belleau et al. Clinical and Molecular Allergy 2005 3:6 doi:10.1186/1476-7961-3-6