Prothrombin Time (PT)

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PT assesses coagulation factors in extrinsic pathway (F VII) and common pathway (F X, F V, prothrombin, and fibrinogen).
 
Principle
Tissue thromboplastin and calcium are added to plasma and clotting time is determined. The test determines the overall efficiency of extrinsic and common pathways.
 
Equipment
(1) Water bath at 37°C
(2) Test tubes (75 × 12 mm)
(3) Stopwatch
 
Reagents
(1) Thromboplastin reagent: This contains tissue factor and phospholipids and is available commercially.
(2) Calcium chloride 0.025 mol/liter.
 
Specimen
Venous blood is collected from antecubital fossa with a plastic, siliconized glass, or polypropylene syringe and a large bore needle (20 or 21 G in adults, 22 or 23 G in infants). Blood should never be collected from indwelling intravenous lines, as these often contain heparin. Glass syringe should not be used for collection since it activates coagulation. The blood is drawn gently but quickly after a single, smooth venepuncture. The needle is detached from the syringe, and the sample is passed gently into the plastic container. After capping the container, the blood and the anticoagulant are mixed immediately by gentle inversion 5 times. The anticoagulant used for coagulation studies is trisodium citrate (3.2%), with anticoagulant to blood proportion being 1:9. Most coagulation studies require platelet poor plasma (PPP). To obtain PPP, blood sample is centrifuged at 3000-4000 revolutions per minute for 15-30 minutes. Coagulation studies are carried out within 2 hours of collection of sample.
 
Method
(1) Deliver 0.1 ml of plasma in a glass test tube kept in water bath at 37°C.
(2) Add 0.1 ml of thromboplastin reagent and mix.
(3) After 1 minute, add 0.1 ml of calcium chloride solution. Immediately start the stopwatch and record the time required for clot formation.
 
Normal Range
11-16 seconds.
 
Causes of prolongation of PT
(1) Treatment with oral anticoagulants
(2) Liver disease
(3) Vitamin K deficiency
(4) Disseminated intravascular coagulation
(5) Inherited deficiency of factors in extrinsic and common pathways.
 
Uses of PT
(1) To monitor patients who are on oral anticoagulant therapy: PT is the standard test for monitoring treatment with oral anticoagulants. Oral anticoagulants inhibit carboxylation of vitamin K-dependent factors (Factors II, VII, IX, and X) and make these factors inactive.
In patients receiving oral anticoagulants, PT should be reported as a ratio of PT of patient to PT of control; it should not be reported as percentage. Various types of thromboplastin reagents obtained from different sources (like ox brain, rabbit brain, or rabbit lung) are available for PT test. These differ in their responsiveness to deficiency of vit. K-dependent factors. Technique of PT is also different in different laboratories. For standardization and to obtain comparable results, it is recommended to report PT (in persons on oral anticoagulants) in the form of an International Normalized Ratio (INR).

INR =  PT of Patient ISI
          PT of Control
 
International Sensitivity Index (ISI) of a particular tissue thromboplastin is derived (by its manufacturer) by comparing it with a reference thromboplastin of known ISI.
INR should be maintained in the therapeutic range for the particular indication (INR of 2.0-3.0 for prophylaxis and treatment of deep venous thrombosis; INR of 2.5-3.5 for mechanical heart valves). Therapeutic range provides adequate anticoagulation for prevention of thrombosis and also checks excess dosage, which will cause bleeding.
(2) To assess liver function: Liver is the site of synthesis of various coagulation factors, including vitamin Kdependent proteins. Therefore PT is a sensitive test for assessment of liver function.
(3) Detection of vitamin K deficiency: PT measures three of the four vitamin K-dependent factors (i.e. II, VII, and X).
(4) To screen for hereditary deficiency of coagulation factors VII, X, V, prothrombin, and fibrinogen.
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Clinical laboratory professional specialized to external quality assessment (proficiency testing) schemes for Laboratory medicine and clinical pathology. Author/Writer/Blogger

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