Topoisomerase II alpha

DNA Topoisomerase II alpha (Topo IIa) is a nucleic enzyme that affects the topological structure of DNA by interacting with the double-helix DNA, thus playing an important role in DNA replication, transcription, recombination, condensation, and segregation. Type II topoisomerases cut both strands of the DNA helix simultaneously in order to change the linking number of the molecule. Topo IIa is essential in the separation of daughter strands at the end of replication. Failure to separate these strands leads to cell death. In cancers, the Topo IIa is highly expressed in highly-proliferating cells.

Topo IIa has been identified by DNA microarray and transcriptional profiling as a gene that is overexpressed in Cervical Carcinomas. The TOP2A gene is approximately 30 kb in size and encodes a 170 kDa protein. Topo IIa protein is expressed in proliferating cells and in numerous human malignant tumors, including colon, gastric and breast cancers, Lymphomas and others. In certain cancers, such as Peripheral Nerve Sheath Tumors, high expression of this protein is also associated with poor patient survival. Type II topoisomerases are the targets for anticancer drugs, such as topoisomerase II inhibitor therapies like the anthracyclines (Doxorubicin and Epirubicin).

Topo IIa is a rabbit monoclonal antibody derived from cell culture supernatant that is concentrated, dialyzed, filter sterilized and diluted in buffer pH 7.5, containing BSA and sodium azide as a preservative.

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1. Cut and mount 3-4 micron formalin-fixed paraffin-embedded tissues on positive charged slides.
2. Air dry for 2 hours at 58° C.
3. Deparaffinize, dehydrate and rehydrate tissues.
4. Subject tissues to heat epitope retrieval using a suitable retrieval solution such as ImmunoDNA Retriever with Citrate (BSB 0020-BSB 0023) or EDTA (BSB 0030-BSB 0033).
5. Any of three heating methods may be used:
   • Electric Pressure Cooker
     Place standoff rack at base of pressure cooker. Add 1-2 inches of distilled water to the pressure cooker and turn heat to high, and incubate for 15 minutes. Open and immediately transfer slides to room temperature.
   • Water Bath Method
     Place tissues/slides in a pre-warmed staining dish or coplin jar containing the ImmunoDNA Retriever with Citrate or EDTA in a water bath set at 95°-99° C. Incubate for 30-60 minutes.
   • Conventional Steamer Method
     Place tissues/slides in a pre-warmed staining dish or coplin jar containing the ImmunoDNA Retriever with Citrate or EDTA in a Steamer, cover and steam for 30-60 minutes.
6. After heat treatment, transfer slides in ImmunoDNA Retriever with Citrate or EDTA to room temperature and let stand for 15-20 minutes.
7. Wash slides with IHC wash buffer or DI water.
8. Continue IHC staining protocol.