Anti-nmt55 p54nrb NONO Antibody Picoband® (monoclonal, 11E2)

Western blot analysis of nmt55 p54nrb using anti-nmt55 p54nrb antibody (M03515).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 50ug of sample under reducing conditions.
Lane 1: human HELA whole cell lysates,
Lane 2: human placenta tissue lysates,
Lane 3: human MCF-7 whole cell lysates,
Lane 4: human A549 whole cell lysates,
Lane 5: human SW620 whole cell lysates,
Lane 6: human PANC-1 whole cell lysates,
Lane 7: human U20S whole cell lysates,
Lane 8: human K562 whole cell lysates.
After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with mouse anti-nmt55 p54nrb antigen affinity purified monoclonal antibody (Catalog # M03515) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-mouse IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1001) with Tanon 5200 system. A specific band was detected for nmt55 p54nrb at approximately 60KD. The expected band size for nmt55 p54nrb is at 60KD.

Flow Cytometry analysis of U20S cells using anti-nmt55 p54nrb antibody (M03515).
Overlay histogram showing U20S cells stained with M03515 (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with mouse anti-nmt55 p54nrb Antibody (M03515,1μg/1x10⁶ cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (BA1126, 5-10μg/1x10⁶ cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1μg/1x10⁶) used under the same conditions. Unlabelled sample (Red line) was also used as a control.

IHC analysis of nmt55 p54nrb using anti-nmt55 p54nrb antibody (M03515).
nmt55 p54nrb was detected in immunocytochemical section of A431 cell. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 1μg/ml mouse anti-nmt55 p54nrb Antibody (M03515) overnight at 4°C. Biotinylated goat anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1021) with DAB as the chromogen.

IF analysis of nmt55 p54nrb using anti-nmt55 p54nrb antibody (M03515).
nmt55 p54nrb was detected in immunocytochemical section of MCF-7 cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (AR0022) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5μg/mL mouse anti-nmt55 p54nrb Antibody (M03515) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Mouse IgG (BA1126) was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.