London Laboratory Services Group

Anti Thrombin 3 Assay

Antithrombin III (ATIII) is one of the most important regulators of the coagulation system. Reduced concentration of ATIII in blood means a great and well established risk for thrombotic complications. ATIII determinations have thus been found to have a prognostic value for thrombosis.
Heterozygous congenital ATIII deficiency with an ATIII level of about 50% of normal activity appears at a frequency of about one in 5000. This is caused either by decreased biosynthesis or a normal concentration of a defective, not fully active protein. Patients with congenital deficiency, constituting only a few percent of all cases of thrombophilia, nearly always suffer from thromboembolic complications when coagulation is triggered and thus have to be specially taken care of.

APC Resistance Assay

The APC resistance phenotype is, in more than 90% of cases, due to a mutation in the Factor V gene, resulting in a replacement of Arg500 (R) with Gin (Q) in the factor V protein. The abnormal Factor V protein is resistant to inactivation by Activated Protein C. This inability of Activated Protein C to inhibit the pro-coagulant proteins (specifically VIIIa and Va) puts the patient at risk of thrombosis. By performing an APTT-based screening assay this abnormality is identified and the specific genetic testing can then performed.

Euglobulin Fibrinolysis Assay

This is a screening test for increased fibrinolytic activity. Fibrinogen, plasminogen, plasminogen activators and plasmin make up the euglobulin fraction isolated from plasma. The activity of this precipitate is directly proportional to the fibrinolytic activity. The plasmin inhibitors are not present in this precipitate and require specific assays for their analysis.

Factor II Assay

Prothrombin is a vitamin K-dependent proenzyme that functions in coagulation. There are two types of this deficiency, a congenital version called hypoprothrombinemia, and an acquired version called dysprothrombinemia.
A life-long bleeding disorder when congenital, factor II deficiency is extremely rare. In fact, only 30 cases of this hereditary clotting factor defect have been identified in the whole world.
Acquired factor II deficiency is more common. It results from vitamin K deficiency, severe liver disease and therapeutic use of anticoagulant drugs. Risk factors for vitamin K deficiency are prolonged use of antibiotics, bile duct obstruction and intestinal malabsorption (inadequate absorption of nutrients from the intestinal tract) disorders. Some newborns are born with vitamin K deficiency.
If the deficiency is caused by liver disease, the outcome depends on control of the liver problem. Vitamin K administration will correct vitamin K deficiency.
Either form may lead to severe bruising, excessive menstrual bleeding, postoperative hemorrhage or occasional muscle bleeds.

Factor V Assay

Factor V deficiency is also known as Owren's disease or parahemophilia. This deficit was identified in Norway in 1943. Since then about 150 cases have been reported, occurring in both men and women. The exact frequency of this rare disorder is unknown, but is estimated to be one per 1 million.
The role of factor V is to accelerate the activity of thrombin. When levels of factor V are low, blood clotting is delayed or it progresses slowly. People with this deficiency may have occasional nosebleeds, excessive menstrual bleeding and bruising; although, many have no symptoms. The first sign of this condition may be bleeding following surgery.
In this disorder, bleeding ranges from mild to severe. The disease is similar to hemophilia, except that bleeding into joints is uncommon. Bleeding can occur almost anywhere in the body, and death from hemorrhage has occurred with this disorder. Excessive bleeding with menstrual periods and postpartum hemorrhage occurs frequently.
A family history of a bleeding disorder is a risk factor. Men and women are affected equally. The probable outcome is good with proper diagnosis and treatment.
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Factor VII Assay

This extremely rare disorder can be inherited or acquired People who take Coumadin, a drug used to inhibit blood clotting, will have an acquired factor VII deficiency.

With this disorder, bleeding can vary from mild to severe within the same person over time. Bleeding doesn't always correspond with the severity of the deficiency shown in blood tests. A history of bleeding may occur in infancy or childhood. Gastrointestinal and central nervous system bleeding can also occur.

This disorder occurs in one in 500,000 males and females. Congenital factor VII deficiency should be distinguished from acquired factor VII deficiency, which may result from liver disease, vitamin K deficiency or other malabsorption conditions.

When levels of the factor are less than 1% of normal, bleeding can be severe. The trauma of birth may cause bleeding in the head of a newborn. Circumcision may cause prolonged bleeding. Children and adults may suffer bleeding from nose, gums or gastrointestinal tract, and women may suffer excessive menstrual bleeding. The probable outcome is good with proper treatment.

Factor VIII Assay

Hemophilia is a bleeding disorder caused by a deficiency in one of the blood clotting factors. Hemophilia A (often called classic hemophilia) accounts for about 80% of all hemophilia cases. It is a deficiency in clotting factor VIII.
Hemophilia A is a hereditary disorder in which the clotting ability of the blood is impaired and excessive bleeding results. Small wounds and punctures are usually not a problem, but uncontrolled internal bleeding can result in pain and swelling and permanent damage, especially to joints and muscles.
Severity of symptoms can vary and severe forms become apparent early on. Prolonged bleeding is the hallmark of hemophilia A and typically occurs when an infant is circumcised. Additional bleeding manifestations make their appearance when the infant becomes mobile. Mild cases may go unnoticed until later in life when there is excessive bleeding and clotting problems in response to surgery or trauma. Internal bleeding may happen anywhere, and bleeding into joints is common.
The incidence of hemophilia A is one in 10,000 live male births. About 17,000 Americans have hemophilia. Women may have it, but it's very rare. With treatment and management, the outcome is good. Most men with hemophilia are able to lead relatively normal lives.

Factor IX Assay

Hemophilia is a bleeding disorder caused by a deficiency in one of the blood clotting factors. Hemophilia B (also called "Christmas disease" after Stephen Christmas, a British boy in the 20th century who was first diagnosed with it) is a deficiency in clotting factor IX.
Hemophilia A is seven times more common than hemophilia B. The incidence of hemophilia B is one in 34,500 men.
Hemophilia B is a hereditary disorder in which the clotting ability of the blood is impaired and prolonged bleeding results. Small wounds and punctures are usually not a problem, but uncontrolled internal bleeding can result in pain, swelling and permanent damage, especially to joints and muscles.
The outcome is good with treatment and management. Most people with hemophilia B are able to lead relatively normal lives.

Factor X Assay

Factor X deficiency ranks similarly to factor II as one of the rarest inherited clotting disorders, with only about 50 reported cases. Factor X is a vitamin K-dependent clotting factor in the blood.It was first discovered in a man named Stuart from the mountains of North Carolina. He was originally thought to have factor VII deficiency, but then a woman named Prower was found to have a similar clotting abnormality, and the new factor X was discovered.
In a factor X disorder, bleeding ranges from mild to severe. Severity ranges on the level of factor X clotting activity. Women may have severe menstrual bleeding and postpartum hemorrhage. The incidence ranges from 1 out of 500,000 to 1 out of 1,000,000 people. Newborns may present with prolonged bleeding after circumcision. People with factor activity that is less than 1% of normal are susceptible to severe bleeding. Those with 10% or more are only mildly effected.
Some cases are due to reduced or absent synthesis of the molecule. In other cases, the number of molecules is normal, but they don't work properly. Several genetic variations of factor X deficiency of varying severity have been described.
Even more uncommon than the congenital form of factor X deficiency is the acquired form. It has occasionally developed in patients with liver disease, vitamin K deficiency, amyloidosis, myeloma, leprosy, mycoplasma infections, various tumors and exposure to the fungicide methylbromide.
Factor X levels have been shown to increase during pregnancy, but aggressive replacement therapy is needed for pregnant women or women with a severe deficiency and a history of early and recurrent miscarriage.

Factor XI Assay

Factor XI deficiency is a very rare bleeding disorder, occurring in an estimated one in 100,000 North Americans. It has more variable bleeding tendencies than hemophilia A and B, and people with the disorder usually don't bleed into joints and muscles, as in hemophilia. The disorder is usually only mild, and is typically provoked by surgery.
Factor XI is a protein that helps blood clot and is activated by thrombin. Deficiencies occur rarely, and in all racial groups, but it is particularly common in Ashkenazi Jews. These are primarily Jews of eastern European ancestry, and the relationship is presumably because intermarriage within this closed group, generation after generation, allowed the defective gene to surface more frequently.

Factor XII Assay

First described in 1955, after routine blood tests before surgery on a man named John Hageman, factor XII deficiency is a very rare bleeding disorder. As a group, Asians are born with the deficiency more than any other ethnic group.
The deficiency is somewhat of a medical mystery. It does not cause abnormal bleeding, even with major surgical procedures. The hallmark of severe factor XII deficiency is prolonged APTT-usually more than 100 seconds-in a patient with no history of bleeding.
The outcome is expected to be good even without treatment

Factor XIII Assay

This is perhaps the rarest of all factor deficiencies, affecting just one in several million. A hallmark of this rare inherited deficiency is poor wound healing and abnormal scar formation. The reason is that factor XIII-fibrin stabilization factor-is necessary for clot formation and wound healing.
Factor XIII is responsible for clot stabilization and cross linking of the fibrin polymer in blood. A clot will form in the absence of factor XIII, but it will be unstable.
Factor XIII deficiency is a severe bleeding disorder usually associated with trauma. Typically, it's discovered in newly-born babies who suffer recurrent bleeding from the umbilical stump and hemorrhages after circumcision.
Severely affected patients have a high incidence of intracranial hemorrhage with little or no trauma. Affected women often experience spontaneous abortion unless treated with plasma replacement therapy. Bleeding in surgery is not excessive, but delayed bleeding can occur.

Inhibitors

Patients who develop an antibody against a specific clotting factor are said to have an acquired factor inhibitor. If a patient develops an antibody against their own clotting factor, the immune system "tags" the clotting factor with the antibody. With the attachment of the antibody, the clotting factor is removed from the blood more rapidly than normal. The liver then makes clotting factor faster, but it still cannot make enough to keep up. Thus, the blood clotting factor level falls below the normal range. This results in an increased tendency to bleed.

Factor IX Inhibitor
Factor V Inhibitor
Factor VIII Inhibitor
Factor VIII Inhibitor
Porcine Factor VIII INH

Fibrinogen Antigenic Assay

Abnormalities of fibrinogen production may be congenital or acquired and, in general, involve either decreased production of a normal molecule (afibrinogenemia, hypofibrinogenemia) or production of an abnormal molecule (dysfibrinogenemia). Congenital abnormalities of fibrinogen production are inherited in an autosomal dominant fashion. Acquired abnormalities may be observed in a variety of clinical conditions such as liver diseases or consumptive coagulation disorders.

Heparin Assay

Two relatively new anticoagulants, low-molecular weight heparin (LMWH) and danaparoid (Orgaran®), when present at therapeutic levels, usually do not significantly prolong the activated partial thromboplastin time (PTT). Therefore, when laboratory tests are used to monitor therapeutic anticoagulant levels of LMWH or danaparoid, antifactor Xa assays are necessary. In addition, in some instances the PTT cannot be used to monitor unfractionated heparin. For example, lupus anticoagulants* or certain factor deficiencies (eg, factor XII deficiencies) may prolong the baseline PTT and/or accentuate the PTT prolongation when heparin is added. In these cases, unfractionated heparin may be monitored with antifactor Xa assays (*Note: if the antifactor Xa assay demonstrates that the heparinized PTT is not affected by the lupus anticoagulant, cautious use of the PTT may be tried in that patient).

Heparin Induced Thrombocytopenia Assay

For many years, heparin, used intravenously, has been an effective anti-coagulant, preventing clotting in patients at risk for developing disorders of clotting such as deep vein thrombosis.
Heparin is a sulfated glycosaminoglycan (GAG) and can naturally be found in human tissues and inflammatory cells such as mast cells. Heparin sulfated is located on the endothelial cells lining the blood vessel wall. Heparin in the serum and heparin sulfate bind antithrombin which is the major inhibitor protein of the coagulation cascade. When binding occurs, the antithrombin protein is altered and this accelerates its inhibition of other serine proteases (factors IXa, Xa, XIa, XIIa, kallikrein, and thrombin).
Heparin can be fractionated into two parts using affinity chromatography with immobolized antithrombin. One fraction, the high-affinity heparin, is responsible for nearly all of the anticoagulant activity. The other fraction, low-affinity heparin, has virtually no anticoagulant activity. The anticoagulant activity is expressed in units relative to an international standard (IU). Low molecular weight heparin (LMWH) is prepared by fractionation.
Patients with this disorder are receiving heparin therapy and develop antibodies to antigens on platelets. This leads to thrombocytopenia which may be profound.

Hypercoagulable Screen

A hypercoagulable disorder is an inherited or acquired condition that increases the risk of inappropriate or excessive thrombosis (blood clot) formation. Clotting is a normal response to blood vessel or tissue injury. When a blood vessel is injured, it begins to leak blood, either externally or into body tissues. The body stops this blood loss through a complex clotting process called hemostasis. During hemostasis, the injured blood vessel constricts to reduce blood flow, platelets adhere to the injury site and clump together to form a loose platelet plug, and the coagulation cascade is initiated. During the cascade process, the body sequentially activates coagulation factors, proteins that create a net of fibrin threads, weave them through the platelet plug and stabilize the resulting blood clot. This clot functions as a barrier to further blood loss, one that stays in place until the injury has healed.
Usually, the body activates the clotting process, regulates its speed and volume with feedback mechanisms, and after the site has healed, breaks down the clot and removes it. Hypercoagulable disorders occur when something goes wrong with the clotting process. If the process activates inappropriately, does not self regulate properly, or resists being broken down, then there can be inappropriate and/or excessive blood clot formation. Blood clots are referred to as thrombi (one - thrombus) when they form in a blood vessel; thrombi may break off and block another blood vessel in another part of the body, where they are referred to as emboli (one - embolus). Thrombi most commonly form in the deep veins of the lower legs (deep venous thrombosis, or DVT), where they may cause pain and swelling; they may also occur in arteries, leading to strokes and heart attacks. Emboli most commonly involve the lungs (pulmonary emboli), where they can cause chest pain and shortness of breath.
Inherited hypercoagulable disorders are relatively rare and are usually due to a genetic mutation that leads to a deficiency or dysfunction in the coagulation protein that the gene produces. They may be seen in heterozygous (one gene copy) or homozygous (two gene copies) form. If someone has two mutated gene copies, they tend to have a more severe form of the condition, and if they are heterozygous in more than one condition, the risk of clotting tends to be additive (and sometimes they multiply the risk). With inherited hypercoagulable disorders the first thrombotic episode may be seen at a relatively young age (less than 40 years of age). The patient may have recurrent thrombosis, a family history of thrombosis, and blood clots in unusual sites (such as cerebral veins, hepatic veins, and renal veins)
Acquired disorders are more common than inherited ones. They may be related to antiphospholipid antibodies, liver disease, or to some cancers. DIC (disseminated intravascular coagulation) is a life threatening acute acquired condition that causes tiny clots throughout the body. It uses up coagulation factors at an accelerated rate, leading to both bleeding and clotting. The next few pages describe several hypercoagulable disorders.

Lupus Anticoagulant

Lupus anticoagulant testing is used to help determine the cause of an unexplained thrombosis, recurrent fetal loss, or a prolonged aPTT test. It is ordered to help determine whether a prolonged aPTT is due to a specific inhibitor (an antibody against a specific coagulation factor), or to a nonspecific inhibitor, like the lupus anticoagulant. It may be ordered along with anticardiolipin antibody and anti-beta2-glycoprotein I assay to check for antiphospholipid syndrome. If someone tests positive for the lupus anticoagulant, the test may be done again in several weeks to see if the antibody was due to a temporary condition or is a chronic issue. Occasionally lupus anticoagulant testing may be ordered to help determine the cause of a positive VDRL/RPR test for syphilis (both anticardiolipin and lupus antibodies will test false positive with these tests).
Because there are other inhibitors and analytical variables that can cause abnormal test results, several different tests are used to confirm the presence of a lupus anticoagulant. Typically these may include: aPTT, prothrombin time (PT), dilute or modified Russell viper venom screen (dRVVT or MRVVT), and a hexagonal (II) phase phospholipid assay (Staclot-LA test) or kaolin clot time. A thrombin time test may also be done to rule out heparin contamination (this is a drug used for anticoagulant therapy), and a fibrinogen test may be done to rule out hypofibrinogenemia. These two conditions can cause prolongations in the test results and interfere with lupus anticoagulant detection.

Plasminogen

Plasminogen is the precursor of plasmin, which lyses fibrin clots. Hereditary plasminogen deficiency is rare, and it may predispose to venous thrombosis.

Platelet Function Screen

Platelets are vital for normal blood clotting. Produced in the bone marrow, they circulate in the blood until they are needed. When there is an injury to a blood vessel, platelets adhere to the injury site (with the help of von Willebrand factor, which acts as the "glue"), aggregate with other platelets, release compounds that stimulate further aggregation, and form a loose platelet plug in a process called primary hemostasis. At the same time, platelets support the coagulation cascade, a series of steps that involves the sequential activation of clotting factors. This secondary hemostasis process culminates in the formation of strands of fibrin that are woven through the loose platelet plug, cross-linked to form a fibrin net, and compressed to form a stable clot that remains in place until the injury has healed. When the clot is no longer needed, other factors break the clot down and remove it.
If there are insufficient platelets, or if they are not functioning normally, a patient may be at an increased risk of excessive bleeding. The number of platelets can be easily determined with a platelet count, but the overall platelet function is more difficult to measure. Unfortunately, there is no one test that identifies all problems with platelet function, nor is there widespread agreement on which test(s) are best for each circumstance.

Protein C Antigen & Protein C Functional

Deficiency of Protein C is associated with recurrent venous thrombosis and pulmonary embolism, especially in young adults. Aquired deficiencies of Protein C are associated with hepatic disorders, oral anticoagulant therapy and disseminated intravascular coagulation.

Protein S Antigen & Protein S Functional

Protein S deficiency may be hereditary or aquired. Acquired deficiency may be observed during pregnancy, oral anticoagulant therapy, oral contraception use, in liver desease, in newborn infants as well as in other clinical conditions. Deficiency of Protein S has been associated with a high risk of developing venous thromboembolism especially in young people.

Reptilase

Clotting time similar to thrombin time except that a snake venom (Reptilase®) is used instead of thrombin. The Reptilase® time is prolonged by decreased or dysfunctional fibrinogen, or high levels of fibrin degradation products (FDP). Dysfibrinogenemia is an uncommon hereditary or acquired condition characterized by dysfunctional fibrinogen.

Thrombin Time

Measures clotting time of the last step in the coagulation cascade, which is the conversion of fibrinogen into fibrin by thrombin. Useful for diagnosis of dysfibrinogenemia. Very sensitive to low amounts of heparin, hirudin, or argatroban anticoagulants.

Tissue Factor

Tissue factor (TF) is a cell membrane-bound glycoprotein (MW 46 kDa) and a member of the class 2 cytokine receptor family. It is composed of a hydrophilic extracellular domain, a membrane-spanning hydrophobic domain, and a cytoplasmic tail of 21 residues, including a non-disulfide-linked cysteine. The mature protein, which is post-translationally modified to include carbohydrate moieties, is biologically active. Upon exposure to blood, perivascular cell-bound TF binds to factor VII, a vitamin K-dependent serine protease unique among coagulation factor zymogens in that it exists in a partially active state. Cleavage of factor VII to VIIa by thrombin, factor IXa, Xa, or XIIa increases its activity 100-fold. The affinity of TF for factor VIIa is increased by anionic phospholipids. The TF-factor VIIa complex can directly or indirectly activate factor X and hence generate thrombin (factor IIa)

Tissue Plasminogen Activator

Naturally occurring substance in the body tissues that activates the enzyme plasmin that is able to dissolve blood clots. Human tPA, produced in bacteria by genetic engineering, has, like streptokinase, been used to dissolve blood clots in the coronary arteries of heart-attack victims. It has been shown to be more effective than streptokinase when used in conjunction with heparin.

von Willebrands Screen

VD is classified into type 1 (the most frequent for being 70 - 80% of VWD), type 2 or 3 (1 to 3% of VWD) groups. Type 1 shows a reduction of VWF although its structure and functionality is normal. In type 3 VWF is almost absent in plasma. In type 2, the quantity of VWF in plasma may be normal or slightly reduced but its molecular structure and its functionality is abnormal.

Type 2 may be further characterized into subtypes by multimeric VWF structure.
Acquired VWD due to either:
1. Autoantibodies
2. Chronic or acute inflammatory diseases or processes involving damage of the vascular endothelium