The McGill Physiology Virtual Lab

Blood Laboratory

Hemostasis > PT and PTT tests
  When a body tissue is injured and begins to bleed, it initiates a sequence of clotting factor activities -the coagulation cascade-  leading to the formation of a blood clot. This cascade is comprised of three pathways: extrinsic, intrinsic, and common.

Two laboratory tests are used commonly to evaluate coagulation disorders: Prothrombin Time (PT) which measures the integrity of the extrinsic system as well as factors common to both systems and Partial Thromboplastin Time (PTT), which measures the integrity of the intrinsic system and the common components.

Blood Clot Formation  
Clotting is a function of plasma.  It depends upon the orderly interaction of a group of plasma proteins (which are sequentially activated following vascular injury) with some phospholipid (from either damaged tissue or platelets) and some Ca++. The final stages include the formation of thrombin, which then converts:

 soluble plasma protein fibrinogen ---------------->insoluble fibrin.

Another factor converts the fibrin into a cross-linked polymer which stabilizes the platelet plug and traps RBCs in the meshwork to form the actual blood clot. Depending on the type of vascular damage or abnormality, clotting can be initiated and proceed according to two different cascading pathways: the intrinsic (initiated by contact with and abnormal/foreign surface) or the extrinsic (initiated by exposure to tissue factors). Note that:

  •  the two pathways converge, so that the final steps are common to the two schemes  
  • although clotting can be initiated via either the more rapid (15-20 secs) extrinsic scheme or the slower (2-6 mins) intrinsic scheme,  
  •  the division into two pathways is only an artifact of  in vitro testing: the two pathways interconnect at several levels. In vivo, both pathways must be activated for effective hemostasis.
  • both coagulation pathways, by a series of feedback mechanisms, control their own activity (e.g. traces of thrombin enhance the activity of earlier factors in the scheme).

Note also that, in addition to the coagulation-promoting factors, there are also substances in blood which inhibit coagulation (e.g., an anti-thrombin factor which inactivates thrombin).  Whether or not blood coagulates depends on the balance that exists between the two groups of factors (pro-coagulants and anti-coagulants).  

Prothrombin time test

 
The PT test is used to monitor patients taking certain medications as well as to help diagnose clotting disorders.

A sample of the patient's blood is obtained by venipuncture. The blood is decalcified (by collecting it into a tube with oxalate or citrate ions) to prevent the clotting process from starting before the test. The blood cells are separated from the liquid part of blood (plasma) by centrifugation. The PT test is performed by adding the patient's plasma to some source of Tissue Factor (e.g.: a protein, thromboplastin, from homogenized brain tissue) that converts prothrombin to thrombin. The mixture is then kept in a warm water bath at 37C for one to two minutes. Calcium chloride (excess quantities of ionized calcium) is added to the mixture in order to counteract the sodium citrate and allow clotting to start. The test is timed from the addition of the calcium chloride until the plasma clots. This time is called the Prothrombin Time.

The prothrombin test specifically evaluates the presence of factors VII, V, and X, prothrombin, and fibrinogen. A prothrombin time within the 11 -15 second range (depends on the source of thromboplastin used) indicates that the patient has normal amounts of the above clotting factors.

A prolonged prothrombin time indicates a deficiency in any of factors VII, X, V, prothrombin, or fibrinogen. It may mean that the patient has a vitamin K deficiency (vitamin K is a co-factor in the synthesis of functional factors II (prothrombin), VII, IX and X) or  a liver disease (the liver is the site of synthesis of the plasma protein factors). The prothrombin time of patients receiving a vitamin K-competing coumarin drug such as warfarin (anticoagulation therapy used in deep venous thrombophlebitis) will also be prolonged, usually in the range of one and one half to two times the normal PT time.

Activated Partial Thromboplastin Time test

 

The activated partial thromboplastin time (aPTT) is a test performed to investigate bleeding disorders and to monitor patients taking an anticlotting drug such as heparin which inhibits factors X and thrombin, while activating anti-thrombin.

The aPTT test uses blood which is decalcified to prevent clotting before the test begins. The plasma is separated by centrifugation. (Ionized) Calcium and activating substances are added to the plasma to start the intrinsic pathway of the coagulation cascade. The substances are: kaolin (hydrated aluminum silicate) and cephalin. Kaolin serves to activate the contact-dependent Factor XII, and cephalin substitutes for platelet phospholipids. The partial thromboplastin time is the time it takes for a clot to form, measured in seconds. Normally, the sample will clot in 35 seconds.

PTT measures the integrity of the intrinsic system (Factors XII, XI, VIII, IX) and common clotting pathways.

Increased levels in a person with a bleeding disorder indicate a clotting factor may be missing or defective. At this point, further investigation is needed and warrants the use of sensitive assays for specific coagulation factors. Liver disease decreases production of factors, increasing the PTT.

To continue to the next section: Blood Cell Indices, click here