Fibrinogen

 
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Fibrinogen concentration can be measured in three ways, the most accurate of which is clottable fibrinogen.

Heat precipitation

Heat precipitation is performed on anticoagulated samples (EDTA preferred). It is mostly used for determination of fibrinogen concentration as an indicator of inflammation in horses, ruminants and camelids and can be included on routine hemograms in horses. Fibrinogen is an acute phase reactant protein and the liver increases production of this protein in response to inflammatory cytokines. It is considered a moderate, but delayed, marker of inflammation in horses. Concentrations increase within 24-48 hours of an inflammatory stimulus and moderate increases are seen. Values can remain increased for a while after resolution of the inflammation. Studies have shown that other acute phase proteins (specifically serum amyloid A in horses, haptoglobin in cattle) and reactants (iron concentrations in serum or plasma) are better markers of acute inflammation. In horses, decreased serum iron concentration and increased serum amyloid A precede increases in fibrinogen and values drop more rapidly with resolution of inflammation. The test also does not appear to be sensitive to inflammation in camelids (anecdotal observations). This method of fibrinogen determination is quite crude (values change in 100 mg/dL quantities and the minimum detectable limit is 100 mg/dL) and the technique is not sensitive enough to detect decreased fibrinogen concentration in coagulation abnormalities. Measurement of clot table fibrinogen is more accurate for both increased or decreased concentrations. Thus, this test should always be used as a crude guide to fibrinogen concentrations (normal values do not rule out inflammation or hypofibrinogenemia). It is not recommended in small animals since clottable fibrinogen measurement is more accurate and readily available.

  • Method of measurement: EDTA-anticoagulated whole blood is drawn into two microhematocrit tubes. One is immediately centrifuged and the total protein determined on the supernatant plasma by refractometer. The second tube is placed in a 56°C water bath for 3 minutes. Fibrinogen is a pyroglobulin and will precipitate when heated. The heated microhematocrit tube is then centrifuged and the total protein determined by refractometer. The difference between the two refractometer measurements is equivalent to the fibrinogen concentration in mg/dL, i.e. the fibrinogen that was removed from the plasma in the second tube by heating and centrifuging. Results are reported in mg/dL.
  • Interpretation: Normal concentrations do not rule out inflammation (values correlate poorly to clottable fibrinogen results). Results will be inaccurate if there are interferents in blood that affect measurement of refractive index (lipemia, hemolysis). High concentrations  are seen in the following conditions:
    • Inflammation
    • Renal disease in cows

Clottable fibrinogen (Clauss method)

This is part of coagulation screening panels in some laboratories and is interpreted along with the prothrombin time (PT), activated partial thromboplastin time (APTT), and thrombin clot time (TCT).

  • Method of measurement: Fibrinogen concentration is determined using a modified TCT, where the time taken for the patient’s sample to clot after adding a standard amount of thrombin is measured. The TCT is modified for fibrinogen measurement by the inclusion of heparin inhibitors and dilution of the patient sample, which minimizes the effects of inhibitors of fibrin polymerization on the clotting time. The clotting time is then compared to a standard curve prepared from (preferably) species-specific fibrinogen. The TCT is inversely proportional to the fibrinogen concentration, therefore a long TCT indicates a hypofibrinogenemia and vice versa. Fibrinogen concentration is reported in milligrams per deciliter (mg/dL).
  • Interpretation: Interferents (lipemia, hemolysis) should not impact the assay unless the TCT is performed using photo-optical measurements (which are affected by sample turbidity).
    • Low fibrinogen (hypofibrinogenemia): This occurs with synthetic liver failure and DIC (latter is due to consumption).
    • High fibrinogen (hyperfibrinogenemia): Fibrinogen is an acute phase reactant protein. Liver synthesis increases in response to inflammatory cytokines, e.g. interleukin-1, interleukin-6. In one prospective study of 116 dogs admitted to an emergency center, fibrinogen concentrations were significantly higher in dogs with localized inflammation (553 ± 270 mg/dL) and systemic inflammatory response syndrome or sepsis (500 ± 234 mg/dL) than clinically healthy blood donor dogs (213 ± 103 mg/dL), although there was overlap between groups. Fibrinogen concentrations showed good discrimination between dogs with localized or systemic inflammation compared to healthy controls, based on receiver operator characteristic curves with area under the curves ranging from 0.868-0.906 (Torrente et al 2015). High fibrinogen concentrations may also be seen in renal disease in cats and cattle (mechanism unknown).

Fibrinogen antigen

Fibrinogen antigen can be measured with clot table fibrinogen to diagnose dysfibrinogenemia, i.e. where fibrinogen is present but cannot be clotted by thrombin, or afibrinogenemia (absence of fibrinogen).

  • Method of measurement: This is done using immunologic assays in which fibrinogen in the patient sample reacts with specific antibody against fibrinogen. Techniques would include ELISA and nephelometry (immunoturbidometric assay). The amount of fibrinogen antigen is then determined by comparing the change in turbidity (nephelometry) or color change (ELISA) to that of a standard curve of serially diluted species-specific (preferably) fibrinogen. Fibrinogen antigen measurement is only offered by specialized veterinary coagulation testing laboratories, such as the Comparative Coagulation Laboratory at Cornell University, where it is done by ELISA.
  • Interpretation: Results are interpreted with clottable fibrinogen results.
    • Low fibrinogen antigen, low clottable fibrinogen: Hypofibrinogenemia (acquired disorders, e.g. synthetic liver failure, DIC), afibrinogenemia (inherited) if no antigen detected with very long TCT.
    • Normal fibrinogen antigen, low clottable fibrinogen: Dysfibrinogenemia (abnormal fibrinogen structure). This could be inherited or acquired (high concentrations of FDPs or paraparoteins).

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