Anti-cIAP2 antibody (ab23423)

ICC/IF image of ab23423 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab23423, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iT^TM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).

HeLa cells were incubated at 37°C for 24h with vehicle control (0 µM) and different concentrations of TMCB (ab120289). Decreased expression of cIAP2 in HeLa cells correlates with an increase in TMCB concentration, as described in literature.

Whole cell lysates were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 40µg of each were loaded on the gel and the WB was run under reducing conditions. After transfer the membrane was blocked for an hour using 5% BSA before being incubated with ab23423 at 2 µg/ml and ab1801 at 2 µg/ml overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP (ab97051 ) at 1/10000 dilution and visualised using ECL development solution.