Chromogenix Coamatic® Protein C is a chromogenic assay kit for the quantitative determination of protein C activity in human citrated plasma.
Protein C plays an important role in the anticoagulant regulatory mechanisms. Hereditary and/or acquired Protein C deficiency has been shown to be a risk factor for development of venous thrombosis.
No influence of heparin levels up to 3 IU/ml. Simple and rapid performance gives optimal user convenience. Excellent accuracy and precision as well as reagent stability. Validated and documented protocols for a wide range of instruments. Suitable for both large and small laboratories.
Reagents, Packaging, Storage and Stability
- S-2366™ 6 mg, 2 vials. Lyophilized chromogenic substrate pyroGlu-Pro-Arg-pNA·HCl. Stability after reconstitution: 3 months at 2-8°C in the original vial.
- Protein C activator 1.2 U, 2 vials. Lyophilized venom enzyme from Southern Copperhead Snake (Agkistrodon Contortrix Contortrix), with bovine serum albumin (stabilizer) and Ciprofloxacin® (preservative). Stability after reconstitution: 3 months at 2-8°C in the original vial.
The sealed reagents are stable at 2-8°C until the expiry date printed on the label. WARNING: Do not use reagents beyond the expiry date printed on the package label. Discard if the substrate solution appears yellow.
- Test tube: 72
- Microplate: 288
- Automated: up to 180
Protein C in plasma is activated by a specific enzyme from Southern Copperhead Snake venom. The amount of activated protein C is determined by the rate of hydrolysis of the chromogenic substrate S-2366™. The pNA release measured at 405 nm is proportional to the Protein C level in the range from 0-120% of normal plasma.
|Protein C activator|
|Protein C||——→||Activated Protein C (APC)|
|S-2366™||——→||Peptide + pNA|
Protein C is a vitamin K dependent plasma protein which plays an important role in the anticoagulant regulatory mechanisms. It circulates as a zymogen and is converted to an active serine protease, activated Protein C (APC), by the action of thrombin in presence of thrombomodulin. APC regulates the coagulation system by proteolytic cleavage and inactivation of activated factors V and VIII. Hereditary and/or acquired Protein C deficiency has been shown to be a risk factor for development of venous thrombosis. Read more…
|What is the Chromogenix Coamatic® Protein C measurement principle?|
|In the Chromogenix Coamatic® Protein C assay, what substances could interfere with the assay, how will they affect results, and what can be done to overcome the interference?|
|A low protein C activity is expected in aprotinin treated patients because aprotinin is an inhibitor of activated protein C. Oral anticoagulant therapy interferes with the formation of g-carboxyglutamic acid moiety of the protein C molecules during biosynthesis in the liver, which results in a loss of anticoagulant activity. Non-carboxylated forms of protein C molecules that are inactive in vivo can still be activated by snake venom or thrombin-thrombomodulin and retain amidolytic activity in vitro. Assays using chromogenic substrates will therefore over-estimate the true level of protein C activity in plasma from patients receiving OAC’s. Streptokinase also influences the hydrolysis of S-2366™. Sample blank activities should be determined with plasma from patients with thrombolytic disorder treated with streptokinase, as well as plasma where contact activation is suspected, should be compared to sample blank activities. A high blank activity may indicate contact activation has occurred. S-2366™ is also sensitive to thrombin. This interference can be quenched by the addition of a thrombin inhibitor such as I-2581.|
|Explain the types of protein C deficiency, and the clinical manifestations.|
|Hereditary protein C deficiency is inherited as an autosomal dominant trait. Heterozygotes for protein C deficiency have protein C activity or antigen levels of 30-70% normal, whereas homozygotes with a severe defect have levels below 1%. The prevalence of protein C deficiency is 2-5% in patients with thromboembolic disease. Two types of protein C deficiency states are recognized. In type I deficiency, which is the most common type of disorder, the plasma concentration of protein C is reduced both in functional and immunological assays. This reflects a genetic defect causing a reduced biosynthesis of protein C. Type II deficiency is characterized by normal protein C antigen levels, but with decreased functional activity. This type of defect reflects synthesis of abnormal molecules with reduced function. The most common clinical manifestation of symptomatic heterozygous protein C deficiency is deep vein thrombosis (DVT) of the lower extremities. Patients with homozygous protein C deficiency usually suffer from severe and fatal thrombosis in the early stage of life.
Protein C deficiency can also be acquired. Protein C level is influenced by various diseases and drugs such as DIC, DVT, liver disease, sepsis, oral anticoagulant therapy, and surgery.
In contrast, elevated Protein C levels have been reported in such cases as diabetic patients and with the use of anabolic steroids and oral contraceptives. Elevated levels of protein C have no known clinical significance.
|What effect does variation of plasma levels of protein C have on the APC ratio?|
|Variations in plasma levels of protein C have no influence on the APC ratio since a standardized amount of exogenous APC is added.|
|Describe the protein C pathway and APC resistance. What is the relative risk of venous thrombosis for those who are APC resistant?|
|Protein C is a vitamin-K-dependent glycoprotein and plasma proenzyme of a serine protease that plays a key role in the down-regulation of blood coagulation. It is activated in vivo by the thrombin-thrombomodulin complex on the surface of intact endothelial cells. Activated protein C (APC) functions as a circulating anticoagulant through proteolytic cleavage and inactivation of the coagulation factors Va and VIIIa. The cleavage occurs at three sites in the heavy chain of each protein. The anticoagulant activity of APC is potentiated by the free form of Protein S (about 60% of PS in plasma is bound to C4bBP, and 40% is in free form) and FV. APC Resistance is actually due to a defect in the protein C pathway, in the factor V molecule as opposed to the activated protein C molecule. APC Resistance is an autosomal dominant hereditary defect mainly due to a point mutation resulting in an amino acid change in the FV gene (Ag506 to Gln mutation, or Factor V Leiden mutation). The mutation destroys one of the three cleavage sites, rendering FVa partially resistant to APC-mediated degradation. APC resistance occurs in 3-5% of the general population, but varies largely in different parts of the world. Up to 90% of APC resistance cases are due to the Factor V:Q506 gene mutation. The relative risk of DVT for carriers of the FV:Q506 mutation is estimated to be 8-fold for heterozygotes and 80-fold for homozygotes.|
|Protein concentrations in plasma|