Factor VIII

Factor VIII (F VIII) is an essential clotting factor. The lack of normal F VIII causes Hemophilia A, an inherited bleeding disorder. FVIII is a glycoprotein procofactor synthesized and released into the bloodstream by the liver. In the circulating blood, it is mainly bound to von Willebrand factor (vWF, also known as Factor VIII-related antigen) to form a stable complex. Upon activation by thrombin or Factor Xa, it dissociates from the complex to interact with Factor IXa, the coagulation cascade. It is a cofactor to Factor IXa in the activation of Factor X, which, in turn, with its cofactor Factor Va, activates more thrombin. Thrombin cleaves fibrinogen into fibrin which polymerizes and crosslinks (using Factor XIII) into a blood clot.

This antibody reacts with endothelial cells in normal, reactive, and neoplastic blood cells. F VIII antibody has helped to establish the endothelial nature of some lesions of disputed histogenesis, e.g., Kaposi`s Sarcoma and Cardiac Myxoma. Not all endothelial cells synthesize (or store) this molecule; therefore, it should not be surprising that not all tumors of endothelial differentiation (benign or malignant) react with this antigen.

Factor VIII antibody is a purified immunoglobulin fraction of rabbit antiserum, diluted in phosphate buffered saline, pH 7.6, with protein, and preserved with sodium azide.

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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.