Tumor necrosis factor alpha (TNF-alpha) is a signaling molecule that interacts with one of two receptors on cells targeted for apoptosis. The apoptotic signal is transduced inside these cells by cytoplasmic adaptor proteins. The protein encoded by this gene is a death domain-containing adaptor protein that interacts with the death domain of TNF-alpha receptor 1 to activate mitogen-activated protein kinase (MAPK) and propagate the apoptotic signal. It is membrane-bound and expressed at a higher level in neoplastic cells than in normal cells. Several transcript variants encoding different isoforms have been described for this gene.

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Flow cytometric analysis of MADD expression in HeLa cells using MADD antibody . Green, isotype control; red, MADD.

Immunocytochemical staining of HeLa cells with MADD antibody . Nuclei were stained blue with DAPI; MADD was stained magenta with Alexa Fluor® 647. Images were taken using Leica stellaris 5. Protein abundance based on laser Intensity and smart gain: Medium. Scale bar, 20 μm.

Western blotting analysis using MADD antibody . Total cell lysates (30 μg) from various cell lines were loaded and separated by SDS-PAGE. The blot was incubated with MADD antibody and HRP-conjugated goat anti-rabbit secondary antibody respectively. Image was developed using FeQ™ ECL Substrate Kit .

Western blotting analysis using MADD antibody . MADD expression in wild type (WT) and MADD shRNA knockdown (KD) HeLa cells with 20 μg of total cell lysates. Hsp90 α serves as a loading control. The blot was incubated with MADD antibody and HRP-conjugated goat anti-rabbit secondary antibody respectively. Image was developed using NaQ™ ECL Substrate Kit .

Validation of MADD knockdown using flow cytometry. Wild-type(WT, Blue) and knockdown(KD, Green) HeLa cells were stained with MADD antibody and analyzed using BD flow cytometer.

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