Factor XI BioAssay(TM) ELISA Kit (Antibodies only)

Factor XI (FXI, plasma thromboplastin antecedent) is a coagulation protein produced in the liver that circulates in plasma at approximately 5 microg/ml (30 nM). The mass of FXI is 160 kDa as determined by SDS-PAGE under non-reducing conditions and 80 kDa upon reduction. FXI consists of two identical 80 kDa subunits linked by disulphide bonds. Each subunit consists of a tandem repeat of four apple domains followed by a serine protease catalytic domain. Cleavage of FXI by activated factor XII or thrombin converts each subunit into a two-chain form and generates two active sites per FXIa molecule. The mass of FXIa is 160 kDa unreduced, but upon reduction FXIa migrates as a heavy chain of 50 kDa and a light chain of 30 kDa. The catalytic site of FXIa resides in the light chain. In plasma, FXI or FXIa circulates in non-covalent 1:1 complex with high molecular weight kininogen, which acts as a cofactor in the activation of FXI by activated factor XII. The activity of FXIa is regulated by platelets and by several proteinase inhibitors including, in order of decreasing importance, C1-inhibitor, a2antiplasmin, a1antitrypsin and antithrombin. Heparin has relatively little effect on the rate of inhibition of FXIa by antithrombin. The only known natural substrate for activated FXI (FXIa) is factor IX (Christmas factor) and the only cofactor required for this reaction is ionized calcium 1-3.\n\nPrinciple of Sandwich-style ELISA\nAffinity-purified antibody to FXI is coated onto the wells of a microtitre plate. Any remaining binding sites on the plastic wells are blocked with an excess of bovine serum albumin. The plates are washed and plasma or other fluids containing FXI are applied. The coated antibody will capture the FXI in the sample. After washing the plate to remove unbound material, a peroxidase conjugated second antibody to FXI is added to the plate to bind to the captured FXI. After washing the plate to remove unbound conjugated antibody, the peroxidase activity is expressed by incubation with o- phenylenediamine (OPD). After a fixed development time the reaction is quenched with the addition of H2SO4 and the colour produced is quantified using a microplate reader. The color generated is proportional to the concentration of FXI present in the sample.\n\nMaterials Supplied:\nAntibodies for 5x96 well plates\n\nKit Components:\n\nF0018-27A: Capture Antibody, 1x500ul\nAffinity-purified sheep anti-FXI IgG suitable for coating ELISA plates.\nYellow capped vial\n\nF0018-27B: Detecting Antibody, 1x500ul\nAffinity-purified sheep anti-FXI labeled with HRP suitable for detection of captured FXI\nRed capped vial\n\nStorage and Stability:\nMay be stored at 4 degrees C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20 degrees C. Aliquots are stable for at least 12 months. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.

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SPECIFICATIONS

Catalog Number

F0018-57

Size

1Kit

Applications

ELISA

Form

Supplied as a liquid in buffer, 50% glycerol.

Purity

Purified by affinity chromotography.

References

1. Wuillemin WA, Minnema M, Meijers JCM, Roem D, Erenberg AJM, Nuijens JH, ten Cate H, Hack EC; Inactivation of Factor XIa in Human Plasma Assessed by Measuring Factor XIa-Protease Inhibitor Complexes: Major Role for C1-Inhibitor. Blood 85:1517, 1995.2. DeLa Cadena R, Watchtfogel YT, Colman RW, in Hemostasis and Thrombosis, 3rd Edition, eds. RW Colman, J Hirsh, VJ Marder and EW Salzman, pp. 219-240, J.B. Lippincott Co., Philadelphia, 1994. 3. Baglia FA, Seaman FS, Walsh, PN; The Apple 1 and 4 domains of Factor XI Act to Synergistically Promote the Surface-Mediated Activation of Factor XI by Factor XIIa. Blood 85:2078, 1995.4. Nix,B, Wild D, in Immunoassays, A Practical Approach, editor J.P. Gosling, pp. 239-261, Oxford University Press, 2000. 5. NCCLS. Evaluation of the Linearity of Quantitative Analytical Methods; Proposed Guidline

Additional Info

Recognizes human Factor XI.