Topoisomerase II alpha/TOP2A Antibody

• Catalog Number: orb570416
• Category: Antibodies
• Description: Topoisomerase II alpha/TOP2A Antibody
• Clonality: Polyclonal
• Species/Host: Rabbit
• Isotype: Rabbit IgG
• Conjugation: Unconjugated
• Reactivity: Human
• Form/Appearance: Lyophilized
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Purification: Immunogen affinity purified.
• Immunogen: A synthetic peptide corresponding to a sequence in the middle region of human Topoisomerase II alpha/TOP2A, identical to the related mouse and rat sequences.
• UniProt ID: P11388
• MW: 174 kDa
• Tested applications: FC, ICC, IF, IHC, WB
• Application notes: Western blot, 0.25-0.5μg/ml, Human Immunohistochemistry (Paraffin-embedded Section), 0.5-1μg/ml, Human 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
• Cross Reactivity: No cross-reactivity with other proteins.
• Antibody Type: Primary Antibody
• 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: DNA topoisomerase 2-alpha; DNA topoisomerase II, a
• Note: For research use only
• Expiration Date: 12 months from date of receipt.

Flow Cytometry analysis of 293T cells using anti-TOP2A antibody. Overlay histogram showing 293T 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-TOP2A 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 TOP2A using anti-TOP2A antibody. TOP2A was detected in immunocytochemical section of U20S cells. 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-TOP2A 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 TOP2A using anti-TOP2A antibody. TOP2A was detected in paraffin-embedded section of human Lung cancer tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 µg/ml rabbit anti-TOP2A 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 TOP2A using anti-TOP2A antibody. TOP2A was detected in paraffin-embedded section of human rectal cancer tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 µg/ml rabbit anti-TOP2A 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 TOP2A using anti-TOP2A 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 50 ug of sample under reducing conditions. Lane 1: human A549 whole cell lysates, Lane 2: human U2OS whole cell lysates, Lane 3: human HEK293 whole cell lysates, Lane 4: human K562 whole cell lysates. 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-TOP2A 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 1:10000 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 TOP2A at approximately 174 KD. The expected band size for TOP2A is at 174 KD.