MG-132, proteasome inhibitor

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - MG-132, proteasome inhibitor (AB141003)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells, treated with 2μM MG-132 (ab141003) for 18 hours or untreated, labeling SQSTM1 / p62 (phospho S349) with ab211324 at 1/100 dilution, followed by Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) secondary antibody at 1/200 dilution (green).

Confocal image showing cytoplasmic staining on HeLa cell line. The expression increased after treatment with 2μM MG-132 (ab141003) for 18 hours.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594)) at 1/200 dilution (red).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor^® 488) (ab150077) at 1/1000 dilution.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - MG-132, proteasome inhibitor (AB141003)

Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cell line from cervix adenocarcinoma) cells, treated with 2μM MG-132 (ab141003) for 18 hours (red) or untreated (green), labeling SQSTM1 / p62 (phospho S349) with ab211324 at 1/500 dilution compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor^® 488) at 1/2000 dilution was used as the secondary antibody.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - MG-132, proteasome inhibitor (AB141003)

ab62352 staining Nrf2 in untreated HeLa cells (top panel) and treated HeLa cells (bottom panel). Cells were treated with 2μM of MG-132 for 18 hours (ab141003). The cells were fixed with 4% paraformaldehyde (10 min) then permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab62352 at 1μg/ml concentration and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at 4°C followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.
Image was taken with a confocal microscope (Leica-Microsystems TCS SP8).

• IP

Lab

Immunoprecipitation - MG-132, proteasome inhibitor (AB141003)

SQSTM1 / p62 (phospho S349) was immunoprecipitated from 0.35 mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate treated with 2μM MG-132 (ab141003) for 18h with ab211324 at 1/30 dilution.

Western blot was performed from the immunoprecipitate using ab211324 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : HeLa treated with 2μM MG-132 (ab141003) for 18h whole cell lysate, 10 μg (Input).
Lane 2 : ab211324 IP in HeLa treated with 2μM MG-132 (ab141003) for 18h whole cell lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab211324 in HeLa treated with 2μM MG-132 (ab141003) for 18h whole cell lysate.

Blocking and dilution buffer : 5% NFDM/TBST.

All lanes:

Immunoprecipitation - Anti-SQSTM1 / p62 (phospho S349) antibody [EPR20451] (<a href='/products/primary-antibodies/sqstm1-p62-phospho-s349-antibody-epr20451-ab211324'>ab211324</a>)

Predicted band size: 47 kDa

false

Exposure time: 10s

• Chemical Structure

Lab

Chemical Structure - MG-132, proteasome inhibitor (AB141003)

2D chemical structure image of ab141003, MG-132, proteasome inhibitor