Anti-hnRNP D/AUF1/HNRNPD Antibody Picoband® (monoclonal, 4F3)

Western blot analysis of hnRNP D/AUF1/HNRNPD using anti-hnRNP D/AUF1/HNRNPD antibody (M09982-1).
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 HEK293 whole cell lysates,
Lane 2: human K562 whole cell lysates,
Lane 3: human A431 whole cell lysates,
Lane 4: human HEPG2 whole cell lysates,
Lane 5: human placenta tissue lysates,
Lane 6: rat brain tissue lysates,
Lane 7: mouse brain tissue 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-hnRNP D/AUF1/HNRNPD antigen affinity purified monoclonal antibody (Catalog # M09982-1) 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 hnRNP D/AUF1/HNRNPD at approximately 43-45KD. The expected band size for hnRNP D/AUF1/HNRNPD is at 38KD.

IHC analysis of hnRNP D/AUF1/HNRNPD using anti-hnRNP D/AUF1/HNRNPD antibody (M09982-1).
hnRNP D/AUF1/HNRNPD was detected in paraffin-embedded section of human rectal cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml mouse anti-hnRNP D/AUF1/HNRNPD Antibody (M09982-1) overnight at 4°C. Biotinylated goat anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1021) with DAB as the chromogen.

IHC analysis of hnRNP D/AUF1/HNRNPD using anti-hnRNP D/AUF1/HNRNPD antibody (M09982-1).
hnRNP D/AUF1/HNRNPD was detected in paraffin-embedded section of rat brain tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml mouse anti-hnRNP D/AUF1/HNRNPD Antibody (M09982-1) overnight at 4°C. Biotinylated goat anti-mouse IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) (Catalog # SA1021) with DAB as the chromogen.

IF analysis of hnRNP D/AUF1/HNRNPD using anti-hnRNP D/AUF1/HNRNPD antibody (M09982-1).
hnRNP D/AUF1/HNRNPD was detected in paraffin-embedded section of human rectal cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 4μg/mL mouse anti-hnRNP D/AUF1/HNRNPD Antibody (M09982-1) overnight at 4°C. Biotin conjugated goat anti-mouse IgG (BA1001) was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using DyLight®488 Conjugated Avidin (BA1128). The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Flow Cytometry analysis of SiHa cells using anti- hnRNP D/AUF1/HNRNPD antibody (M09982-1).
Overlay histogram showing SiHa cells stained with M09982-1 (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-hnRNP D/AUF1/HNRNPD Antibody (M09982-1, 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 without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

Flow Cytometry analysis of U937 cells using anti- hnRNP D/AUF1/HNRNPD antibody (M09982-1).
Overlay histogram showing U937 cells stained with M09982-1 (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-hnRNP D/AUF1/HNRNPD Antibody (M09982-1, 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 without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

IF analysis of hnRNP D/AUF1/HNRNPD using anti-hnRNP D/AUF1/HNRNPD antibody (M09982-1).
hnRNP D/AUF1/HNRNPD was detected in an immunocytochemical section of A431 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 5 μg/mL mouse anti-hnRNP D/AUF1/HNRNPD Antibody (M09982-1) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Mouse IgG (BA1126) was used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. Visualize using a fluorescence microscope and filter sets appropriate for the label used.