Figure 3

Inhibition of the basal carbonic anhydrase IX (CA IX) shedding with BB-94 metalloprotease inhibitor. (A) CGL3 cells were biotinylated and then allowed to shed CA IX ECD in the presence and absence of 1 μ M BB-94. The media (m) were collected, the cells were extracted (ex) and the corresponding aliquots of the materials were analysed by immunoprecipitation–immunoblotting as described in Figure 1c. (B) ELISA was used to determine the inhibition of CA IX shedding with 1 μ M BB-94. The cells were treated for 24 h throughout their incubation in normoxia or hypoxia. Extent of inhibition was illustrated as a ratio of the absorbance values of the materials from BB-94-treated cells vs their nontreated counterparts normalised according to total protein concentrations. (C) ELISA assessment of a BB-94 concentration-dependent inhibition of CA IX ECD release in MDCK+CA IX cells. The results are expressed as a ratio between the BB-94-inhibited and control samples. ^*P<0.01 and ^**P<0.001 by Student's t-test.