GammaCoeur™ CVD Risk ELISA Test System

Intended Use

The gamma-prime Fibrinogen assay is intended to be used as part of a risk assessment in adults suspected of/at risk for cardiovascular disease (CVD), for example, adults with hypertension, increased BMI, diabetes, history of smoking, or family history of CVD. It is a semi-automated quantitative ELISA method that uses human venous plasma samples and is performed in clinical laboratories.

Significance and Background

Fibrinogen is a heterogeneous mixture of isoforms with varying relative proportions. Alternative mRNA processing and posttranslational modifications give rise to several different fibrinogen isoforms with widely varying characteristics. In addition, because fibrinogen is a 6-chain molecule containing 2 copies each of the Aα, Bβ, and γ chains, various combinations of altered chains can be assembled, particularly in fibrinogens resulting from heterozygous polymorphisms or mutations. The fibrinogen γ chain has 2 isoforms, the gamma A (γA or simply γ) isoform and the gamma-prime (or γB) isoform. The gamma-prime isoform arises from alternative mRNA processing that results in the substitution of the carboxyl terminal 4 amino acids with a different 20-amino acid sequence. The gamma-prime chain is usually paired with the more common γA chain.

Gamma-prime fibrinogen typically constitutes approximately 10% of total fibrinogen in plasma, although this percentage can vary among individuals. Gamma-prime fibrinogen has several biochemical and biophysical properties that distinguish it from the more common γA isoform. Clots made from fibrinogen containing gamma-prime chains in the presence of factor XIII are highly resistant to fibrinolysis. In addition, the gamma-prime chain contains a binding site for thrombin, and clots made from gamma-prime fibrinogen have been reported to have an altered clot architecture.

Possibly as a result of these properties, recent studies suggest that gamma-prime fibrinogen is a risk factor for cardiovascular disease. An association has been found between gamma-prime fibrinogen concentrations and prevalent coronary artery disease, myocardial infarction, stroke, and inflammation.

Principle of the Test

The GammaCoeur™ assay is a two-site sandwich immunoassay that measures the gamma-prime fibrinogen isoform in human plasma samples. Gamma-prime fibrinogen in the sample binds to a monoclonal capture antibody coated on the assay plate microwells. After incubation, excess sample (including unbound non-gamma-prime fibrinogen [i.e., gammaA/gammaA fibrinogen] and other plasma components) are removed by washing. Enzyme-labeled polyclonal anti-fibrinogen detection antibody is added, which binds to the captured gamma-prime fibrinogen, forming a “capture antibody/analyte/detection antibody” sandwich. After another incubation, excess detection antibody is removed by washing. Finally, a substrate for the enzyme-labeled detector is added to generate a colored product. After a last incubation, a stop solution is added to each well and the absorbance of the colored end-product is read on a spectrophotometer. The amount of colored reaction product formed is proportional to the amount of enzyme-labeled antibody bound to analyte, which is in turn proportional to the amount of gamma-prime fibrinogen present in the sample. All reagents required are provided in the GammaCoeur^™ Test Kit.

Benefits and Features

• System utilizes gamma prime fibrinogen
• Gamma prime fibrinogen forms clots resistant to fibrinolysis
• Independently associated with CVD
• Framingham study shows patients in top 1/3 gamma prime fibrinogen with 3x odds of heart attack
• CE Marked