Anti-UBE1C/UBA3 Antibody

• Catalog Number: orb334570
• Category: Antibodies
• Description: Anti-UBE1C/UBA3 Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: FC, ICC, IF, IHC, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Rabbit IgG
• Immunogen: A synthetic peptide corresponding to a sequence at the C-terminus of human UBE1C, identical to the related mouse and rat sequences.
• Antibody Type: Primary Antibody
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 52 kDa
• UniProt ID: Q8TBC4
• Storage: Maintain refrigerated at 2-8°C for up to 2 weeks. For long term storage store at -20°C in small aliquots to prevent freeze-thaw cycles.
• Alternative names: NEDD8-activating enzyme E1 catalytic subunit; 6.3.
• Note: For research use only
• Application notes: Western blot, 0.1-0.5μg/ml, Human, Rat Immunohistochemistry (Paraffin-embedded Section), 0.5-1μg/ml, Mouse, Rat Immunocytochemistry/Immunofluorescence, 2μg/ml, Human Flow Cytometry (Fixed), 1-3μg/1x106 cells, Human. Add 0.2ml of distilled water will yield a concentration of 500ug/ml
• Expiration Date: 12 months from date of receipt.

Flow Cytometry analysis of A549 cells using anti-UBE1C antibody. Overlay histogram showing A549 cells (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 rabbit anti-UBE1C Antibody (1 µg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (5-10 µg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1 µg/1x10^6) 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 UBE1C using anti-UBE1C antibody. UBE1C was detected in immunocytochemical section of A431 cell. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 2 µg/mL rabbit anti-UBE1C Antibody overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG 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.

IHC analysis of UBE1C using anti-UBE1C antibody. UBE1C was detected in a paraffin-embedded section of mouse testis 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 1 µg/ml rabbit anti-UBE1C Antibody overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.

IHC analysis of UBE1C using anti-UBE1C antibody. UBE1C was detected in a paraffin-embedded section of rat testis 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 1 µg/ml rabbit anti-UBE1C Antibody overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.

Western blot analysis of UBE1C using anti-UBE1C antibody. 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 30 ug of sample under reducing conditions. Lane 1: human Hela whole cell lysate, Lane 2: human HEK293 whole cell lysate, Lane 3: rat brain tissue lysate. After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-UBE1C antigen affinity purified polyclonal antibody 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-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit with Tanon 5200 system. A specific band was detected for UBE1C at approximately 52 kDa. The expected band size for UBE1C is at 52 kDa.