Cytokeratin HMW/AE3

Cytokeratins are intermediate-filament keratins found in the intracytoplasmic cytoskeleton of epithelial tissue. There are two types of cytokeratins: the low-weight, acidic Type I cytokeratins and the high-weight, basic or neutral Type II cytokeratins. Cytokeratins are usually found in pairs comprising a Type I cytokeratin and a Type II cytokeratin. Expression of these cytokeratins is frequently organ or tissue-specific.

Cytokeratin, High Molecular Weight AE3 (HMW, CK 8) is capable of recognizing all basic cytokeratins; therefore, it is a broadly reactive antibody staining most epithelia and their neoplasms. Cytokeratin HMW/AE3 stains normal and neoplastic cells of epithelial origin. CK HMW is primarily found in the non-squamous epithelia and is present in the majority of Adenocarcinomas and Ductal Carcinomas. It is absent in Squamous Cell Carcinomas. Hepatocellular Carcinomas are defined by the use of antibodies that recognize only cytokeratin 8 and 18.

Cytokeratin HMW AE3 is a mouse 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.