Anti Rabbit Tissue Factor

Description
Tissue Factor (TF) is a 45 kDa transmembrane cell surface glycoproteinknown for its role in initiating coagulation. It is comprised of threedomains: an extracellular domain (aa 1-219), a hydrophilic spanningdomain (aa 220-242) and a cytoplasmic tail (aa 243-263). Released intothe blood stream following disruption of the endothelium, tissue factorfunctions as a receptor and cofactor for factors VII and VIIa. Contactbetween TF and blood is sufficient to initiate the extrinsic pathway ofcoagulation. This initiation requires the participation of a series ofmolecules; factor VII, factor X or factor IX, charged phospholipids andcalcium ions, and their ability to complex with tissue factor. TheTF/FVIIa complex efficiently activates both factor X and factor IX, thusinitiating both the intrinsic and extrinsic coagulation pathways.

No. 4511 is highly inhibitory of rabbit tissue factor procoagulantactivity (rabbit brain thromboplastin) in a single-stage prothrombinassay. It shows no effect on human tissue factor. While its epitopespecificity is undetermined, its reactivity toward rabbit tissue factorin Western blot under non-reducing conditions and reduced reactivitytoward rabbit tissue factor under reducing conditions suggests that theantibody recognizes an epitope that includes disulfide bonds.

Preparation
No. 4511 is a murine IgG1 monoclonal antibody purified from ascitesfluid via Protein G affinity chromatography. Immunoaffinity purifiedrabbit tissue factor (Mr = 45,000) was used as the immunizing antigen.

Presentation
Screw-capped glass vial containing 100 μg of purified IgG1 lyophilizedfrom a 200 μL solution of 0.15M Phosphate Buffered Saline, 100 mMMannitol, pH 7.4.

Reconstitution
Add 200 μL of filtered deionized or sterile water to generate a 0.5mg/mL stock solution.

Storage
Store lyophilized antibody at 2°-8°C. Aliquot and store reconstitutedantibody at –20°C or colder.

Applications
A. Western Blot Analysis
No. 4511 at a primary antibody concentration of 5 μg/mL detects aslittle as 5 ng or native and recombinant rabbit tissue factor (ADI No.4520) under non-reducing conditions. Multiple bands may be visualizeddue to variations in glycosylation.

B. Immunohistochemistry
No. 4511 has been used for localizing rabbit tissue factor in theadventitia of blood vessels and in normal and diseased rabbit glomeruli(paraffin-embedded tissue sections). A primary antibody concentration of20 μg/mL in conjunction with Bouin’s fixation is recommended.1

C. Immunoprecipitation/Affinity Purification
No. 4511 is suitable for these applications. A 5:1 molar ration ofantibody:protein is recommended for immunoprecipitation.

D. Inhibition
5 μg of No. 4511 inhibits 99% of the the procoagulant activity ofrabbit tissue factor. It demonstrates no inhibitory effects toward humantissue factor.

References
1. Tipping, P. G., Erlich, J. H., Apostolopoulos, J., Mackman, N.,Loskutoff, D. and Holdsworth, S. R. Glomerular tissue factor expressionin crescentic glomerulonephritis. Correlations beween antigen, activity,and mRNA. American Journal of Pathology 1995, 147(6): 1736-1748.